Myoglobin facilitates angiotensin II-induced constriction of renal afferent arterioles.

Vasoconstriction plays an important role in the development of acute kidney injury in rhabdomyolysis. We hypothesized that myoglobin enhances the angiotensin II (ANG II) response in afferent arterioles by increasing superoxide and reducing nitric oxide (NO) bioavailability. Afferent arterioles of C57Bl6 mice were isolated perfused, and vasoreactivity was analyzed using video microscopy. NO bioavailability, superoxide concentration in the vessel wall, and changes in cytosolic calcium were measured using fluorescence techniques. Myoglobin treatment (10-5 M) did not change the basal arteriolar diameter during a 20-min period compared with control conditions. NG-nitro-l-arginine methyl ester (l-NAME, 10-4 M) and l-NAME + myoglobin reduced diameters to 94.7 and 97.9% of the initial diameter, respectively. Myoglobin or l-NAME enhanced the ANG II-induced constriction of arterioles compared with control (36.6 and 34.2%, respectively, vs. 65.9%). Norepinephrine responses were not influenced by myoglobin. Combined application of myoglobin and l-NAME further facilitated the ANG II response (7.0%). Myoglobin or l-NAME decreased the NO-related fluorescence in arterioles similarly. Myoglobin enhanced the superoxide-related fluorescence, and tempol prevented this enhancement. Tempol also partly prevented the myoglobin effect on the ANG II response. Myoglobin increased the fura 2 fluorescence ratio (cytosolic calcium) during ANG II application (10-12 to 10-6 M). The results suggest that the enhanced afferent arteriolar reactivity to ANG II is mainly due to a myoglobin-induced increase in superoxide and associated reduction in the NO bioavailability. Signaling pathways for the augmented ANG II response include enhanced cytosolic calcium transients. In conclusion, myoglobin may contribute to the afferent arteriolar vasoconstriction in this rhabdomyolysis model.

[Psychological treatments for pain in cancer patients : A systematic review on the current state of research]. / Schmerzpsychologische Interventionen bei onkologischen Patienten : Ein systematisches Review zum aktuellen Stand der Forschung.

BACKGROUND: In cancer patients, pain is one of the main symptoms and especially in the late stages of disease, these symptoms can be associated with considerable suffering. In psycho-oncology, preliminary psychological therapies targeting cancer pain have been tested; however, a systematic review of available interventions is lacking, especially considering their dissemination, evidence base, study quality, and the comparison with established treatments. Therefore, the aim of the current study is to systematically review the current research on psychological treatments for pain in cancer patients. MATERIALS AND METHODS: During May 2014, MEDLINE, PsycINFO, PSYNDEX, and CENTRAL databases were searched. Psychological treatments for pain in adult cancer patients studied in randomized, controlled trials (RCTs) and referring to pain as primary or secondary outcome were included. After examination for inclusion, structured data extraction and assessment followed. Data were synthesized narratively. RESULTS: In the review, 32 RCTs were included. Studies mainly referred to patients with breast cancer or patients in earlier stages of the disease. The methodological quality of included studies was heterogeneous. Most commonly, short interventions were delivered by nurses in out-patient settings. Interventions including education and relaxation techniques were utilized most often, followed by interventions with behavioral or cognitive components. CONCLUSION: A need for research persists regarding efficacy of current psychotherapeutic interventions, or the role of mediator variables (e. g., coping) on pain perception in cancer patients. Studies with high methodological quality which comprehensively and transparently report on interventions and designs are lacking.

Affective disturbances modulate the neural processing of visceral pain stimuli in irritable bowel syndrome: an fMRI study.

OBJECTIVE: To address the role of anxiety and depression symptoms in altered pain processing in irritable bowel syndrome (IBS). DESIGN: In this functional magnetic resonance imaging study, the blood oxygen level-dependent (BOLD) response to rectal distensions delivered at previously determined individual discomfort thresholds was assessed. PATIENTS: 15 female patients with irritable bowel syndrome (IBS) and with normal rectal pain thresholds, and 12 healthy women. MEASURES: The correlation of anxiety and depression symptoms, measured with the Hospital Anxiety and Depression Scale (HADS), with subjective pain ratings and the BOLD response during distension-induced brain activation were analysed within IBS. Group differences in pain-induced brain activation with and without controlling for HADS scores were evaluated. RESULTS: Patients with IBS experienced significantly more pain and discomfort upon rectal distensions in the scanner, despite unaltered rectal sensory thresholds. Anxiety and depression scores were associated with these subjective stimulus ratings, but not with rectal sensory thresholds. Anxiety symptoms in IBS were significantly associated with pain-induced activation of the anterior midcingulate cortex and pregenual anterior cingulate cortex. Depression scores correlated with activation of the prefrontal cortex (PFC) and cerebellar areas within IBS. Group comparisons with the two-sample t test revealed significant activation in the IBS versus controls contrast in the anterior insular cortex and PFC. Inclusion of anxiety and depression scores, respectively, as confounding variables led to a loss of significant group differences. CONCLUSIONS: Altered central processing of visceral stimuli in IBS is at least in part mediated by symptoms of anxiety and depression, which may modulate the affective-motivational aspects of the pain response.

Diabetes and radiocontrast media increase endothelin converting enzyme-1 in the kidney.

Plasma endothelin-1 levels rise in diabetes and after exposure to contrast media suggesting a role in progressive diabetic and acute radiocontrast nephropathies. Here we studied individual and combined effects of streptozotocin-induced diabetes and contrast media on renal endothelin converting enzyme-1 levels in the rat. In vivo, medullary (but not cortical) endothelin converting enzyme protein gradually increased 4 to 5-fold following the induction of diabetes or after the administration of contrast media but rose 15-fold when diabetic rats were given contrast media. Changes in mRNA expression paralleled those of the protein. Immunohistochemistry confirmed that increased tubular and endothelial cell endothelin converting enzyme-1 were most pronounced in the medulla. In vitro, endothelin-1 levels increased 3-fold following incubation of endothelial cells with media high in glucose or with contrast and 4-fold with their combination. Endothelin converting enzyme-1 protein and mRNA expression changed in a similar pattern while prepro endothelin-1 mRNA increased with each insult but not in an additive way. Our study shows that diabetes and contrast media up-regulate renal medullary endothelin converting enzyme-1 expression and synthesis.

Cyclosporine A induces endothelin-converting enzyme-1: Studies in vivo and in vitro.

AIM: Cyclosporine A (CsA) induces renal vasoconstriction and hypoxia and enhances the expression of endothelin-1 (ET-1) pro-hormone (pre-pro-ET-1), plausibly leading to a feed-forward loop of renal vasoconstriction, hypoxia and enhanced synthesis of the potent vasoconstrictor ET-1. Endothelin-converting enzyme (ECE)-1 cleaves big endothelin to generate endothelin (ET)-1 and is upregulated by hypoxia via hypoxia-inducible factor (HIF). We hypothesized that in addition to the direct induction of ET-1 synthesis, CsA might also intensify renal ECE-1 expression, thus contributing to enhanced ET-1 synthesis following CsA. METHODS: CsA was administered to Sprague Dawley rats (120 mg/kg/SC) for 4 days, and renal HIF and ECE-1 expression were assessed with Western blots and immunostaining. Human umbilical vein endothelial cells (HUVEC) and proximal tubular cell line (HK-2) were subjected to CsA, and ECE-1 induction was evaluated using real-time mRNA PCR and Western blots. RESULTS: Cyclosporine A intensified renal parenchymal ECE-1 expression in the rat kidney, particularly in distal nephron segments, along with renal hypoxia (detected by pimonidazole adducts) and HIF expression, in line with our recent observations showing episodic hypoxia in mice subjected to CsA. Furthermore, in cultured normoxic HUVEC and HK-2 cells, CsA dose-dependently induced both pre-pro-ET-1 and ECE-1 mRNA and protein expression, with enhanced ET-1 generation. CONCLUSION: CsA induces ECE-1 via both hypoxic and non-hypoxic pathways. ECE-1 may contribute to increased renal ET-1 generation following CsA, participating in a feed-forward loop of renal parenchymal hypoxia and ET synthesis.

A dual role of miR-22 in rhabdomyolysis-induced acute kidney injury.

AIM: In acute kidney injury (AKI), regions of the kidney are hypoxic. However, for reasons yet unknown, adaptation to hypoxia through hypoxia-inducible factor (HIF) is limited. Here, we studied miR-22, a potential HIF repressor, in normal kidneys, as well as in rhabdomyolysis-induced AKI, a condition where miR-22 is up-regulated. METHODS: AKI in mice was provoked by IM injection of glycerol. Tissue homogenates were processed to determine the levels of candidate RNAs and proteins, as well as global gene expression profiles. Reporter assays quantified in vitro miR-22 activity and its modulation by mimic or inhibitor molecules, under normoxia or hypoxia (1% O2 ) respectively. In vivo, anti-miR-22 molecules were applied to normal mice or prior to induction of AKI. Renal outcome was assessed by measuring plasma creatinine, plasma urea and the levels of the injury markers Kim-1 and Ngal. RESULTS: Renal miR-22 is inducible by hypoxia and represses hypoxia-inducible factor (HIF). Specific inhibition of miR-22 regulates 1913 gene transcripts in kidneys controls and 3386 in AKI, many of which are involved in development or carcinogenesis. Specific inhibition of miR-22 up-regulates tissue protective HIF target genes, yet renal function and injury markers are unchanged or worsened. CONCLUSIONS: miR-22 is a HIF repressor constitutively expressed in the adult kidney and up-regulated in AKI. Specific inhibition of miR-22 is efficient in vivo and profoundly affects renal gene expression in health and disease, including up-regulation of HIF. However, the net effect on rhabdomyolysis-induced AKI outcome is neutral or even negative.

Microbial pathogenesis: lipid rafts as pathogen portals.

The route of initial entry influences how host cells respond to intracellular pathogens. Recent studies have demonstrated that a wide variety of pathogens target lipid microdomains in host cell membranes, known as lipid rafts, to enter host cells as an infectious strategy.

Cyclosporin a induces renal episodic hypoxia.

AIM: Cyclosporin A (CsA) causes renal toxicity. The underlying mechanisms are incompletely understood, but may involve renal hypoxia and hypoxia-inducible factors (Hifs). We sought for hypoxia and Hif in mouse kidneys with CsA-induced toxicity, assessed their time course, Hif-mediated responses and the impact of interventional Hif upregulation. METHODS: Mice received CsA or its solvent cremophore for up to 6 weeks. Low salt diet (Na+ ↓) was given in combination with CsA to enhance toxicity. We assessed fine morphology, renal function, blood oxygen level-dependent magnetic resonance imaging under room air and following changes in breathing gas composition which correlate with vascular reactivity, pimonidazole adducts (which indicate O2 tensions below 10 mmHg), Hif-α proteins, as well as expression of Hif target genes. Stable Hif upregulation was achieved by inducible, Pax8-rtTA-based knockout of von Hippel-Lindau protein (Vhl-KO), which is crucial for Hif-α degradation. RESULTS: Cyclosporin A transiently increased renal deoxyhaemoglobin (R2*). Augmented vascular reactivity was observed at 2 h, but decreased at 24 h after CsA treatment. Na+ ↓/CsA provoked chronic renal failure with tubular degeneration and interstitial fibrosis. Nephron segments at risk for injury accumulated pimonidazole adducts, as well as Hif-α proteins. Remarkably, Hif target gene expression remained unchanged, while factor-inhibiting Hif (Fih) was enhanced. Na+ ↓/CsA/Vhl-KO aggravated morpho-functional outcome of chronic renal CsA toxicity. CONCLUSIONS: Cyclosporin A provokes episodic hypoxia in nephron segments most susceptible to chronic CsA toxicity. Fih is upregulated and likely blocks further Hif activity. Continuous tubular Hif upregulation via Vhl-KO worsens the outcome of chronic CsA-induced renal toxicity.

Adaptive response to hypoxia and remote ischaemia pre-conditioning: a new hypoxia-inducible factors era in clinical medicine.

Transient ischaemia leads to tolerance to subsequent protracted ischaemia. This 'ischaemia pre-conditioning' results from the induction of numerous protective genes, involved in cell metabolism, proliferation and survival, in antioxidant capacity, angiogenesis, vascular tone and erythropoiesis. Hypoxia-inducible factors (HIF) play a pivotal role in this transcriptional adaptive response. HIF prolyl hydroxylases (PHDs), serving as oxygen sensors, control HIFα degradation. HIF-mediated ischaemic pre-conditioning can be achieved with the administration of PHD inhibitors, with the attenuation of organ injury under various hypoxic and toxic insults. Clinical trials are currently under way, evaluating PHD inhibitors as inducers of erythropoietin. Once their safety is established, their potential use might be further tested in clinical trials in various forms of acute ischaemic and toxic organ damage. Repeated transient limb ischaemia was also found to attenuate ischaemic injury in remote organs. This 'remote ischaemic pre-conditioning' phenomenon (RIP) has been extensively studied recently in small clinical trials, preceding, or in parallel with an abrupt insult, such as myocardial infarction, cardiac surgery or radiocontrast administration. Initial results are promising, suggesting organ protection. Large-scale multi-centre studies are currently under way, evaluating the protective potential of RIP in cardiac surgery, in the management of myocardial infarction and in organ transplantation. The mechanisms of organ protection provided by RIP are poorly understood, but HIF seemingly play a role as well. Thus, Inhibition of HIF degradation with PHD inhibitors, as well as RIP (in part through HIF), might develop into novel clinical interventions in organ protection in the near future.

Gene array technology to determine host responses to Salmonella.

Gene expression array technology is a powerful new tool that has already been used to expand our understanding of host-pathogen interactions. There has been a rapid increase in published reports describing use of this approach to profile host responses to pathogenic bacteria and viruses. The large number of array studies currently in progress coupled with increasing accessibility of this new technology promises a plethora of gene expression data on host response to infection in the near future. Recent insights into macrophage and epithelial cell responses to Salmonella infection garnered from array studies are outlined and used as a basis to discuss various future research directions using gene arrays that will advance the field of cellular microbiology. There is an exciting potential for the gene expression data generated in such studies to provide insights into host physiology, the pathophysiology of disease and novel therapeutics.

CSF SNAP-25 in schizophrenia and bipolar illness. A pilot study.

Research efforts to identify and understand the pathophysiology of schizophrenia and bipolar illness are limited by the inability to study neuronal tissue of living patients. An alternative to sampling brain tissue from living patients is to measure neuronal proteins found in cerebral spinal fluid. One such candidate protein is synaptosomal-associated protein 25kDa. Our hypothesis is that the level of this protein in cerebral spinal fluid may be a marker of neuronal pathology. Cerebral spinal fluid from headache, schizophrenic, bipolar, and control subjects was used to measure the SNAP-25 level by quantitative dot blotting. Schizophrenic subjects had significantly elevated levels of SNAP-25 as compared to headache and control subjects. However, there was no significant difference between the bipolar group and schizophrenic or control groups. This study reports on a potentially useful clinical marker in schizophrenia, and the presence of elevated cerebral spinal fluid SNAP-25 may indicate alterations in neuronal functioning.

Pilot study to evaluate a water displacement technique to compare effects of diuretics and ACE inhibitors to alleviate lower extremity edema due to dihydropyridine calcium antagonists.

Combination therapy is required in many patients to achieve goal blood pressure (BP). Calcium antagonists are highly effective antihypertensive drugs in a broad range of demographic groups. Yet, higher doses are associated with an increased frequency of lower extremity edema. The purpose of our open label, single-center clinical trial was to evaluate the use of concomitant pharmacologic therapies to attenuate the lower extremity edema associated with dihydropyridine calcium antagonists therapy using a water displacement technique. Forty-seven patients received 5 mg/day of oral amlodipine for a period of 6 weeks after a 4-week wash-out off of all antihypertensive medications to establish baseline BP. They were then randomized to receive either an additional 5 mg of amlodipine, 25 mg of hydrochlorothiazide (HCTZ), or 20 mg of benazepril for an additional 6 weeks. Blood pressure determinations and water displacement measurements were obtained at the end of the 4-week placebo wash-out period, after 6 weeks of 5 mg/day of oral amlodipine therapy, and after an additional 6 weeks of 5 mg of amlodipine and randomized drug therapy. Adjusted BP reductions (based on pretreatment BP) were -6.8/-3.8 mm Hg for the 10-mg amlodipine group, -9.9/-8.2 mm Hg for the amlodipine (5 mg)/HCTZ (25 mg) group, and -26.2/-16.4 mm Hg for the amlodipine (5 mg)/benazepril (20 mg) group (P < .0167, group 3 v group 1 diastolic BP, which was statistically significant by the improved Bonferroni method). Seventeen of the 47 patients developed at least a 10% increase in lower extremity edema water displacement in response to 5 mg/day of oral amlodipine therapy (36.2%). Adding 5 mg of amlodipine to a baseline of 5 mg of amlodipine resulted in no net change in lower extremity edema (+58.0 mL,+ 0.6% change, n=5). Adding 25 mg of HCTZ reduced lower extremity edema by a mean of 136.3 mL (-11.1% change, n=4). Benazepril reduced water displacement by 204.4 mL (-14.3% change, n=8). Our pilot study indicates that adding an angiotensin converting enzyme inhibitor to a dihydropyridine calcium channel blocker is the most effective way to not only reduce systolic and diastolic BP but also attenuate lower extremity edema. Due to the inherent daily variability of lower extremity edema, power calculations indicate many patients (n=702, 356 in each group) would be needed to compare the antiedema efficacy of the angiotensin converting enzyme inhibitor and the thiazide diuretic.

Massive elevation of procalcitonin plasma levels in the absence of infection in kidney transplant patients treated with pan-T-cell antibodies.

OBJECTIVE: To determine the value of procalcitonin (PCT) monitoring in transplant patients receiving pan-T-cell antibody therapy. DESIGN: Retrospective clinical study. SETTING: A collaborative study between the Institute of Medical Immunology, the Department of Nephrology and Internal Intensive Care, both Charite, Humboldt University Berlin, and the Department of Laboratory Medicine, Friedrichshain Hospital, Berlin, Germany. PATIENTS AND INTERVENTIONS: Thirty-one patients were included in the study: 8 kidney transplant patients with acute rejection episodes, 5 receiving OKT3 monoclonal antibody therapy, 3 receiving steroid bolus therapy; 21 patients undergoing renal transplantation, 11 receiving ATG perioperatively, 10 without ATG administration; 2 patients undergoing renal transplantation and receiving anti-IL-2R mAb. MEASUREMENTS AND RESULTS: Procalcitonin (PCT) and tumor necrosis factor (TNF) alpha plasma levels were measured in infection-free transplant patients treated with the pan-T-cell antibodies ATG or OKT3. We found PCT plasma concentrations up to 600 ng/ml (reference < 0.5 ng/ ml), which are comparable to those seen in severe sepsis. Increases in TNF-alpha plasma levels preceded the rises in PCT. After peaking on day 1 of therapy the PCT plasma concentrations returned to normal values independently of further antibody administration. In contrast, steroid bolus therapy or anti-interleukin 2 receptor mAb administration did not increase plasma PCT or TNF-alpha levels. CONCLUSIONS: PCT monitoring for evaluating infectious complications in kidney transplant patients must be very careful during pan-T-cell antibody therapy.

Measuring synaptosomal associated protein-25 kDa in human cerebral spinal fluid.

Changes in the quantity and distribution of neuronal proteins have been reported in psychiatric and neurological illnesses. The majority of this work has been performed in post-mortem samples and the results are difficult to apply to clinical care. The objective of this study is to develop a methodology that can identify trace amounts of brain proteins in cerebral spinal fluid (CSF). Human cerebral spinal fluid was processed to remove albumin and immunoglobulins. CSF samples were analyzed on Western blots using a monoclonal antibody against SNAP-25. These samples were compared to SNAP-25 immunoprecipitated from CSF, rat and human brain homogenates. The monoclonal antibody Mab 331 identified a single band of 25 kDa in all samples. These results demonstrate that the presynaptic protein SNAP-25 can be identified and measured in CSF.

Action of hypoxia-inducible factor in liver and kidney from mice with Pax8-rtTA-based deletion of von Hippel-Lindau protein.

AIM: Von Hippel-Lindau protein (VHL) provides the degradation of hypoxia-inducible factor (HIF). Tetracycline-induced, Pax8-rtTA-based knockout of VHL (VHL-KO) affects all renal tubules and periportal hepatocytes and leads to sustained upregulation of HIF. Here, we study the phenotype of VHL-KO in both organs, the time course of changes, and long-term morpho-functional outcome. METHODS: Mice with doxycycline-induced VHL-KO and controls (CON) were followed for up to 9 months. Systemic and tissue parameters were evaluated using clinical chemistry, histology, immunohistochemistry, RT-PCR and in situ hybridisation. RESULTS: At day 3 following VHL-KO, substantial abundance of HIF-1α and -2α was detected in the nuclei of hepatocytes and renal tubular epithelia. Hypoxia, induced by bleeding anaemia, did not further augment HIF signal. Erythropoietin mRNA was detectable in hepatocytes but not in the kidney. Vascular endothelial growth factor mRNA was upregulated in kidney but not in liver. At day 7 following VHL-KO, the renal capillary density was enhanced, reaching its maximum at day 14. Blood haemoglobin increased constantly up to day 28 (23.3 vs. 15.8 g dL(-1) , VHL-KO vs. CON). Thereafter, it was kept within the normal range by weekly blood collections. Pathological changes were absent from kidney and liver 9 months after VHL-KO. CONCLUSIONS: Inducible, Pax8-rtTA-based deletion of VHL leads to organ-specific expression of epithelial HIF and erythropoietin in liver and kidney without causing pathological changes. Uniform, maximal and sustained HIF activation along the renal tubule may serve to study the potential benefits of hypoxia adaptation in experimental renal injury.

Chronic activation of vasopressin V2 receptor signalling lowers renal medullary oxygen levels in rats.

AIM: In the present study, we aimed to elucidate the effects of chronic vasopressin administration on renal medullary oxygen levels. METHODS: Adult Sprague Dawley or vasopressin-deficient Brattleboro rats were treated with the vasopressin V2 receptor agonist, desmopressin (5 ng/h; 3d), or its vehicle via osmotic minipumps. Immunostaining for pimonidazole and the transcription factor HIF-1α (hypoxia-inducible factor-1α) were used to identify hypoxic areas. Activation of HIF-target gene expression following desmopressin treatment was studied by microarray analysis. RESULTS: Pimonidazole staining was detected in the outer and inner medulla of desmopressin-treated rats, whereas staining in control animals was weak or absent. HIF-1α immunostaining demonstrated nuclear accumulation in the papilla of desmopressin-treated animals, whereas no staining was observed in the controls. Gene expression analysis revealed significant enrichment of HIF-target genes in the group of desmopressin-regulated gene products (P = 2.6*10(-21) ). Regulated products included insulin-like growth factor binding proteins 1 and 3, angiopoietin 2, fibronectin, cathepsin D, hexokinase 2 and cyclooxygenase 2. CONCLUSION: Our results demonstrate that an activation of the renal urine concentrating mechanism by desmopressin causes renal medullary hypoxia and an upregulation of hypoxia-inducible gene expression.

Behavioural and neural correlates of visceral pain sensitivity in healthy men and women: does sex matter?

INTRODUCTION: We assessed sex differences in behavioural and neural responses to rectal pain stimuli in healthy subjects. METHODS: In age- and body mass index-matched healthy subjects (n = 15 men, 15 women), rectal sensory and pain thresholds were assessed with a pressure-controlled barostat device. The blood oxygen level-dependent response during cued anticipation and painful stimulation was measured using functional magnetic resonance imaging (fMRI). Retrospective pain evaluations were accomplished with visual analogue scales. For fMRI data, region-of-interest (ROI) analyses and additional whole-brain analyses were carried out. RESULTS: There were no sex differences in rectal thresholds or pain ratings. ROI analyses revealed comparable distension-induced activation of the thalamus, somatosensory cortex, insula and dorsolateral prefrontal cortex (DLPFC). Only in additional whole-brain analyses did we find increased activation in women in DLPFC and middle temporal gyrus during pain anticipation and in the cerebellum and medial frontal gyrus during pain. A significant inverse association between rectal pain threshold and distension-induced activation in virtually all ROIs was found in women. In men, pain thresholds and insula activation were positively correlated, as were pain ratings and anterior cingulate cortex activation. CONCLUSIONS: Healthy men and women do not differ in behavioural measures of visceral pain sensitivity. The pattern of neural activation is comparable in the majority of pain-processing brain regions, although women may differ in the activation of DLPFC which could reflect sex differences in cognitive-emotional pain regulation. Women with lower pain thresholds showed greater neural responses, which may be relevant in the pathophysiology of visceral hyperalgesia.