Vasculitides associated with IgG antineutrophil cytoplasmic autoantibodies in childhood

Immunoglobulin (Ig)G antineutrophil cytoplasmic autoantibodies are causally associated with necrotizing vasculitides that are characterized immunopathologically by little or no deposition of immunoreactants, such as Wegener granulomatosis, microscopic polyangiitis, Churg-Strauss angiitis, "renal-limited" vasculitis and a number of drug-induced vasculitides. Clinical routine testing targets the antigens myeloperoxidase and proteinase 3. However, in all of the conditions mentioned, the renal histopathologic findings are indistinguishable. Churg-Strauss angiitis (characterized by necrotizing vasculitis, granulomatous inflammation and tissue eosinophilia), Wegener granulomatosis (characterized by necrotizing vasculitis and granulomatous inflammation) and microscopic polyangiitis (characterized by necrotizing vasculitis) often present with fever, weight loss and a multisystem involvement (ear, nose, throat, lung, eyes, peripheral nerve and heart). Fifty years ago these conditions were very often fatal within 6 months of diagnosis. The introduction of corticosteroids and cyclophosphamide has resulted in a dramatic clinical benefit. Patients who develop treatment-related morbidity can be switched from cyclophosphamide to azathioprine after achieving remission. In patients with less severe disease, methotrexate achieves remission with a success rate similar to that of cyclophosphamide. Plasma exchange, in association with immunosuppression, is likely to be a beneficial therapy for patients with severe kidney disease or pulmonary hemorrhage.

Stereoselective Inhibition of the hERG1 Potassium Channel

A growing number of drugs have been shown to prolong cardiac repolarization, predisposing individuals to life-threatening ventricular arrhythmias known as Torsades de Pointes. Most of these drugs are known to interfere with the human ether à-gogo related gene 1 (hERG1) channel, whose current is one of the main determinants of action potential duration. Prolonged repolarization is reflected by lengthening of the QT interval of the electrocardiogram, as seen in the suitably named drug-induced long QT syndrome. Chirality (presence of an asymmetric atom) is a common feature of marketed drugs, which can therefore exist in at least two enantiomers with distinct three-dimensional structures and possibly distinct biological fates. Both the pharmacokinetic and pharmacodynamic properties can differ between enantiomers, as well as also between individuals who take the drug due to metabolic polymorphisms. Despite the large number of reports about drugs reducing the hERG1 current, potential stereoselective contributions have only been scarcely investigated. In this review, we present a non-exhaustive list of clinically important molecules which display chiral toxicity that may be related to hERG1-blocking properties. We particularly focus on methadone cardiotoxicity, which illustrates the importance of the stereoselective effect of drug chirality as well as individual variations resulting from pharmacogenetics. Furthermore, it seems likely that, during drug development, consideration of chirality in lead optimization and systematic assessment of the hERG1 current block with all enantiomers could contribute to the reduction of the risk of drug-induced LQTS.

A new model of interactive effects of alcohol and high-fat diet on hepatic fibrosis

Alcoholic steatohepatitis (ASH) and nonalcoholic steatohepatitis (NASH) are the most frequent conditions leading to elevated liver enzymes and liver cirrhosis, respectively, in the Western world. However, despite strong epidemiological evidence for combined effects on the progression of liver injury, the mutual interaction of the pathophysiological mechanisms is incompletely understood. The aim of this study was to establish and analyze an experimental murine model, where we combined chronic alcohol administration with a NASH-inducing high-fat (HF) diet.

Evolution of gene expression changes in newborn rats after mechanical ventilation with reversible intubation

Mechanical ventilation (MV) is life-saving but potentially harmful for lungs of premature infants. So far, animal models dealt with the acute impact of MV on immature lungs, but less with its delayed effects. We used a newborn rodent model including non-surgical and therefore reversible intubation with moderate ventilation and hypothesized that there might be distinct gene expression patterns after a ventilation-free recovery period compared to acute effects directly after MV. Newborn rat pups were subjected to 8 hr of MV with 60% oxygen (O(2)), 24 hr after injection of lipopolysaccharide (LPS), intended to create a low inflammatory background as often recognized in preterm infants. Animals were separated in controls (CTRL), LPS injection (LPS), or full intervention with LPS and MV with 60% O(2) (LPS + MV + O(2)). Lungs were recovered either directly following (T:0 hr) or 48 hr after MV (T:48 hr). Histologically, signs of ventilator-induced lung injury (VILI) were observed in LPS + MV + O(2) lungs at T:0 hr, while changes appeared similar to those known from patients with chronic lung disease (CLD) with fewer albeit larger gas exchange units, at T:48 hr. At T:0 hr, LPS + MV + O(2) increased gene expression of pro-inflammatory MIP-2. In parallel anti-inflammatory IL-1Ra gene expression was increased in LPS and LPS + MV + O(2) groups. At T:48 hr, pro- and anti-inflammatory genes had returned to their basal expression. MMP-2 gene expression was decreased in LPS and LPS + MV + O(2) groups at T:0 hr, but no longer at T:48 hr. MMP-9 gene expression levels were unchanged directly after MV. However, at T:48 hr, gene and protein expression increased in LPS + MV + O(2) group. In conclusion, this study demonstrates the feasibility of delayed outcome measurements after a ventilation-free period in newborn rats and may help to further understand the time-course of molecular changes following MV. The differences obtained from the two time points could be interpreted as an initial transitory increase of inflammation and a delayed impact of the intervention on structure-related genes.

Pharmacological interaction of drugs with immune receptors: the p-i concept

Drug-induced hypersensitivity reactions have been explained by the hapten concept, according to which a small chemical compound is too small to be recognized by the immune system. Only after covalently binding to an endogenous protein the immune system reacts to this so called hapten-carrier complex, as the larger molecule (protein) is modified, and thus immunogenic for B and T cells. Consequently, a B and T cell immune response might develop to the drug with very heterogeneous clinical manifestations. In recent years, however, evidence has become stronger that not all drugs need to bind covalently to the MHC-peptide complex in order to trigger an immune response. Rather, some drugs may bind directly and reversibly to immune receptors like the major histocompatibility complex (MHC) or the T cell receptor (TCR), thereby stimulating the cells similar to a pharmacological activation of other receptors. This concept has been termed pharmacological interaction with immune receptors the (p-i) concept. While the exact mechanism is still a matter of debate, non-covalent drug presentation clearly leads to the activation of drug-specific T cells as documented for various drugs (lidocaine, sulfamethoxazole (SMX), lamotrigine, carbamazepine, p-phenylendiamine, etc.). In some patients with drug hypersensitivity, such a response may occur within hours even upon the first exposure to the drug. Thus, the reaction to the drug may not be due to a classical, primary response, but rather be mediated by stimulating existing, pre-activated, peptide-specific T cells that are cross specific for the drug. In this way, certain drugs may circumvent the checkpoints for immune activation imposed by the classical antigen processing and presentation mechanisms, which may help to explain the peculiar nature of many drug hypersensitivity reactions.

Oral imatinib treatment reduces early fibrogenesis but does not prevent progression in the long term

BACKGROUND/AIMS: Transactivated hepatic stellate cells (HSCs) represent the key source of extra cellular matrix (ECM) in fibrotic liver. Imatinib, a potent inhibitor of the PDGF receptor tyrosine kinase, reduces HSC proliferation and fibrogenesis when treatment is initiated before fibrosis has developed. We tested the antifibrotic potential of imatinib in ongoing liver injury and in established fibrosis. METHODS: BDL-rats were gavage fed with 20 mg/kg/d imatinib either early (days 0-21) or late (days 22-35) after BDL. Untreated BDL-rats served as controls. ECM and activated HSCs were quantified by morphometry. Tissue activity of MMP-2 was determined by gelatin zymography. mRNA expression of TIMP-1 and procollagen alpha1(I) were measured by RT-PCR. Liver tissue concentration of imatinib was measured by tandem mass spectrometry. RESULTS: Early imatinib reduced ECM formation by 30% (P=0.0455) but left numbers of activated HSCs and procollagen I expression unchanged. MMP-2 activity and TIMP-1 expression were reduced by 50%. Late imatinib treatment did not alter histological or molecular markers of fibrogenesis despite high imatinib tissue levels. CONCLUSIONS: The antifibrotic effectiveness of imatinib is limited to the early phase of fibrogenesis. In ongoing liver injury other mediators most likely compensate for the inhibited PDGF effect.

A case report of Clarkson's disease: If you don't know it, you'll miss it

The systemic capillary leak syndrome (SCLS), also known as Clarkson's disease, is a rare disorder characterized by paroxysmal capillary hyperpermeability with a shift of plasma fluid from the intravascular to the interstitial space. A 35-year-old previously healthy woman was admitted with rapidly developing hypovolemic shock syndrome, rhabdomyolysis, and diffuse edema. Laboratory analysis revealed a severe hemoconcentration, renal insufficiency, and paraproteinemia. After exclusion of infection, allergy, burning or drug-induced conditions, the clinical presentation was consistent with the diagnosis of SCLS. Though this is a rare entity, the substantial morbidity and mortality associated with it necessitate the physician's awareness in order to provide timely therapy. This report is meant to enhance awareness of SCLS.

BOLD-MRI for the assessment of renal oxygenation in humans: acute effect of nephrotoxic xenobiotics

Hypoxia of renal medulla is a key factor implicated in the development of drug-induced renal failure. Drugs are known to influence renal hemodynamics and, subsequently, affect renal tissue oxygenation. Changes in renal oxygenation can be assessed non-invasively in humans using blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI). This study was designed to test the acute effects of administration of specific drugs in healthy human kidney oxygenation using BOLD-MRI. Acute changes in renal tissue oxygenation induced by the non-steroidal anti-inflammatory drug indomethacin, the iodinated radio-contrast media (RCM) iopromidum, and the calcineurin inhibitors cyclosporine micro-emulsion (CsA-ME) and tracrolimus were studied in 30 healthy volunteers. A modified Multi Echo Data Image Combination sequence was used to acquire 12 T(2)(*)-weighted images. Four coronal slices were selected to cover both kidneys. The mean R(2)(*) (1/T(2)(*)) values determined in medulla and cortex showed no significant changes induced by indomethacin and tacrolimus administration. CsA-ME decreased medullary (P=0.008) and cortical (P=0.004) R(2)(*) values 2 h after ingestion. Iopromidum caused a significant increase in medullary R(2)(*) within the first 20 min after injection (P<0.001), whereas no relevant changes were observed in renal cortex. None of the measurements showed left-right kidney differences. Significant differences in renal medullary oxygenation were evidenced between female and male subjects (P=0.013). BOLD-MRI was efficient to show effects of specific drugs in healthy renal tissue. Cyclosporine increased renal medullary oxygenation 2 h after ingestion of a single dose, whereas indomethacin and tacrolimus showed no effect on renal oxygenation. Injection of iodinated RCM decreased renal medullary oxygenation.

Melarsoprol- and pentamidine-resistant Trypanosoma brucei rhodesiense populations and their cross-resistance

Resistance to melarsoprol and pentamidine was induced in bloodstream-form Trypanosoma brucei rhodesiense STIB 900 in vitro, and drug sensitivity was determined for melarsoprol, pentamidine and furamidine. The resistant populations were also inoculated into immunosuppressed mice to verify infectivity and to monitor whether rodent passage selects for clones with altered drug sensitivity. After proliferation in the mouse, trypanosomes were isolated and their IC(50) values to the three drugs were determined. To assess the stability of drug-induced resistance, drug pressure was ceased for 2 months and the drug sensitivity was determined again. Resistance was stable, with a few exceptions that are discussed. Drug IC(50)s indicated cross-resistance among all drugs, but to varying extents: resistance of the melarsoprol-selected and pentamidine-selected trypanosomes to pentamidine was the same, but the pentamidine-selected trypanosome population showed lower resistance to melarsoprol than the melarsoprol-selected trypanosomes. Interestingly, both resistant populations revealed the same intermediate cross-resistance to furamidine. Resistant trypanosome populations were characterised by molecular means, referring to the status of the TbAT1 gene. The melarsoprol-selected population apparently had lost TbAT1, whereas in the pentamidine-selected trypanosome population it was still present.

Neurotoxicity of glia activated by gram-positive bacterial products depends on nitric oxide production

The present study examined the mechanism by which bacterial cell walls from two gram-positive meningeal pathogens, Streptococcus pneumoniae and the group B streptococcus, induced neuronal injury in primary cultures of rat brain cells. Cell walls from both organisms produced cellular injury to similar degrees in pure astrocyte cultures but not in pure neuronal cultures. Cell walls also induced nitric oxide production in cultures of astrocytes or microglia. When neurons were cultured together with astrocytes or microglia, the cell walls of both organisms became toxic to neurons. L-NAME, a nitric oxide synthase inhibitor, protected neurons from cell wall-induced toxicity in mixed cultures with glia, as did dexamethasone. In contrast, an excitatory amino acid antagonist (MK801) had no effect. Low concentrations of cell walls from either gram-positive pathogen added together with the excitatory amino acid glutamate resulted in synergistic neurotoxicity that was inhibited by L-NAME. The induction of nitric oxide production and neurotoxicity by cell walls was independent of the presence of serum, whereas endotoxin exhibited these effects only in the presence of serum. We conclude that gram-positive cell walls can cause toxicity in neurons by inducing the production of nitric oxide in astrocytes and microglia.

Targeted cell replacement with bone marrow cells for airway epithelial regeneration

It has been suggested that some adult bone marrow cells (BMC) can localize to the lung and develop tissue-specific characteristics including those of pulmonary epithelial cells. Here, we show that the combination of mild airway injury (naphthalene-induced) as a conditioning regimen to direct the site of BMC localization and transtracheal delivery of short-term cultured BMC enhances airway localization and adoption of an epithelial-like phenotype. Confocal analysis of airway and alveolar-localized BMC (fluorescently labeled) with epithelial markers shows expression of the pulmonary epithelial proteins, Clara cell secretory protein, and surfactant protein C. To confirm epithelial gene expression by BMC, we generated transgenic mice expressing green fluorescent protein (GFP) driven by the epithelial-specific cytokeratin-18 promoter and injected BMC from these mice transtracheally into wild-type recipients after naphthalene-induced airway injury. BMC retention in the lung was observed for at least 120 days following cell delivery with increasing GFP transgene expression over time. Some BMC cultured in vitro over time also expressed GFP transgene, suggesting epithelial transdifferentiation of the BMC. The results indicate that targeted delivery of BMC can promote airway regeneration.

Concomitant injuries are an important determinant of outcome of high-grade blunt hepatic trauma

BACKGROUND: Little is known about the clinical importance of concomitant injuries in polytraumatized patients with high-grade blunt liver injury. A retrospective single-centre study was performed to investigate the safety of non-operative management of liver injury and the impact of concomitant intra- and extra-abdominal injuries on clinical outcome. METHODS: Some 183 patients with blunt liver injury were admitted to Berne University Hospital, Switzerland, between January 2000 and December 2006. Grade 3-5 injuries were considered to be high grade. RESULTS: Immediate laparotomy was required by 35 patients (19.1 per cent), owing to extrahepatic intra-abdominal injury (splenic and vascular injuries, perforations) in 21 cases. The mortality rate was 16.9 per cent; 22 of the 31 deaths were due to concomitant lesions. Of 81 patients with high-grade liver injury, 63 (78 per cent) were managed without surgery; liver-related and extra-abdominal complication rates in these patients were 11 and 17 per cent respectively. Grades 4 and 5 liver injury were associated with hepatic-related and extra-abdominal complications. CONCLUSION: Concomitant injuries are a major determinant of outcome in patients with blunt hepatic injury and should be given high priority by trauma surgeons. An algorithm for the management of blunt liver injury is proposed.

New perspectives in long QT syndrome

Long QT Syndrome (LQTS) is a cardiac channelopathy characterized by prolonged ventricular repolarization and increased risk to sudden death secondary to ventricular dysrrhythmias. Was the first cardiac channelopathy described and is probably the best understood. After a decade of the sentinel identification of ion channel mutation in LQTS, genotype-phenotype correlations have been developed along with important improvement in risk stratification and genetic guided-treatment. Genetic screening has shown that LQTS is more frequent than expected and interestingly, ethnic specific polymorphism conferring increased susceptibility to drug induced QT prolongation and torsades de pointes have been identified. A better understanding of ventricular arrhythmias as an adverse effect of ion channel binding drugs, allow the development of more safety formulas and better control of this public health problem. Progress in understanding the molecular basis of LQTS has been remarkable; eight different genes have been identified, however still 25% of patients remain genotype-negative. This article is an overview of the main LQTS knowledge developed during the last years.

Targeted gene transfer of hepatocyte growth factor to alveolar type II epithelial cells reduces lung fibrosis in rats

Inefficient alveolar wound repair contributes to the development of pulmonary fibrosis. Hepatocyte growth factor (HGF) is a potent growth factor for alveolar type II epithelial cells (AECII) and may improve repair and reduce fibrosis. We studied whether targeted gene transfer of HGF specifically to AECII improves lung fibrosis in bleomycin-induced lung fibrosis. A plasmid encoding human HGF expressed from the human surfactant protein C promoter (pSpC-hHGF) was designed, and extracorporeal electroporation-mediated gene transfer of HGF specifically to AECII was performed 7 days after bleomycin-induced lung injury in the rat. Animals were killed 7 days after hHGF gene transfer. Electroporation-mediated HGF gene transfer resulted in HGF expression specifically in AECII at biologically relevant levels. HGF gene transfer reduced pulmonary fibrosis as assessed by histology, hydroxyproline determination, and design-based stereology compared with controls. Our results indicate that the antifibrotic effect of HGF is due in part to a reduction of transforming growth factor-β(1), modulation of the epithelial-mesenchymal transition, and reduction of extravascular fibrin deposition. We conclude that targeted HGF gene transfer specifically to AECII decreases bleomycin-induced lung fibrosis and may therefore represent a novel cell-specific gene transfer technology to treat pulmonary fibrosis.

Characterization of choline uptake in Trypanosoma brucei and involvement of a mitochondrial carrier in resistance to choline analogs

Choline is an essential nutrient for eukaryotic cells, where it is used as precursor for the synthesis of choline-­containing phospholipids, such as phosphatidylcholine (PC). Our experiments showed – for the first time – that Trypanosoma brucei, the causative agent of human African sleeping sickness, is able to take up choline from the culture medium to use for PC synthesis, indicating that trypanosomes express a transporter for choline at the plasma membrane. Further characterization in procyclic and bloodstream forms revealed that choline uptake is saturable and can be inhibited by HC-3, a known inhibitor of choline uptake in mammalian cells. To obtain additional insights on choline uptake and metabolism, we investigated the effects of choline-analogs that were previously shown to be toxic for T. brucei parasites in culture. Interestingly, we found that all analogs tested effectively inhibited choline uptake into both bloodstream and procyclic form parasites. Subsequently, selected compounds were used to search for possible candidate genes encoding choline transporters in T. brucei, using an RNAi library in bloodstream forms. We identified a protein belonging to the mitochondrial carrier family, previously annotated as TbMCP14, as prime candidate. Down‐regulation of TbMCP14 by RNAi prevented drug-­induced loss of mitochondrial membrane potential and conferred 8­‐fold resistance of T. brucei bloodstream forms to choline analogs. Conversely, over‐expression of the carrier increased parasite susceptibility more than 13-­fold. However, subsequent experiments demonstrated that TbMCP14 was not involved in metabolism of choline. Instead, growth curves in glucose‐depleted medium using RNAi or knock‐out parasites suggested that TbMCP14 is involved in metabolism of amino acids for energy production. Together, our data demonstrate that the identified member of the mitochondrial carrier family is involved in drug uptake into the mitochondrion and has a vital function in energy production in T. brucei.