Myoinhibitory peptides in the brain of the cockroach Leucophaea maderae and colocalization with pigment-dispersing factor in circadian pacemaker cells

Myoinhibitory peptides (MIPs) are a family of insect W(X(6))Wamides with inhibitory effects on visceral muscles and juvenile hormone synthesis. Although MIPs are widely distributed within the nervous system, a detailed analysis of their distribution and function in insect brains is still missing. We analyzed the distribution of MIPs in the brain of the cockroach Leucophaea maderae. We focused on the accessory medulla (AMe), a small neuropil near the medulla that acts as the master circadian clock. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) and Nano-LC electrospray ionization (ESI) mass spectrometry revealed five Lem-MIPs in preparations of the AMe and corpora cardiaca. The complete sequences of two of these peptides were identified. Immunocytochemistry revealed wide distribution of MIP-related peptides in the cockroach brain. The superior median protocerebrum, parts of the central complex, and the tritocerebrum showed particularly dense immunostaining. In contrast, only a few local interneurons were stained in the antennal lobe and a few extrinsic neurons in the mushroom body, including a giant neuron innervating the calyces. The noduli of the AMe showed dense immunostaining, and neurons in all AMe cell groups except the anterior neurons were labeled. Pigment-dispersing factor- (PDF) and MIP immunostaining was colocalized in two neurons of the AMe. No colocalization of MIP- and PDF immunostaining was detected in the anterior optic commissure, but two small PDF-immunoreactive commissural fibers near the posterior optic commissure showed colocalized MIP immunostaining. The results suggest that several MIPs participate in different functional circuits of the circadian system and are involved in multiple brain circuits of the Madeira cockroach.     

Neuronal damage after moderate hypoxia and erythropoietin

Both mild hypoxia and exogenous erythropoietin may protect the brain against subsequent severe hypoxia, and the conditioning effect of transient hypoxia is partly mediated by hypoxia-induced endogenous erythropoietin. We now observed in several experimental models that combining transient hypoxia and exogenous erythropoietin may cause neuronal damage. High-dose erythropoietin (40 IU/ml) profoundly impeded synaptic transmission of rat hippocampal slice cultures when used in conjunction with moderate hypoxia (10% O2 for two 8-h periods). Addition of erythropoietin increased viability of cultured rat embryonic cortical neurons at 21% O2 but decreased viability under hypoxic conditions (2% O2) in a dose-dependent fashion. Death of human neuronal precursor cells challenged by oxygen and glucose deprivation was increased by erythropoietin when cells were cultured under hypoxic but not under normoxic conditions. In neonatal rats exposed to moderate hypoxia plus erythropoietin, numbers of degenerating cerebral neurons were increased, as compared to controls or rats subjected to either hypoxia or erythropoietin alone. Thus, erythropoietin may aggravate rather than ameliorate neuronal damage when administered during transient hypoxia.     

Ostreolysin induces sustained contraction of porcine coronary arteries and endothelial dysfunction in middle- and large-sized vessels

Ostreolysin (Oly), a cytolytic and cardiotoxic protein from the oyster mushroom (Pleurotus ostreatus), is lethal for mice with an LD₅₀ of 1170 μg/kg following intravenous application. Its cardiotoxicity is associated with hyperkalemia, which is probably a consequence of potassium released from the lysed cells. Moreover, sub-micromolar concentrations of Oly induce a concentration-dependent increase in rat aortic ring tension, suggesting that ischaemia, and consequent hypoxic injury of cardiomyocytes, could also derive from vasospasm induced by this toxic protein.The purpose of the present study was to demonstrate histopathological lesions caused by Oly after parenteral application to rats, and to define the mechanisms of Oly-induced vasoconstriction using inhibitors verapamil, lanthanum chloride, and selective endothelin receptor antagonist TBC3214, which have different molecular targets, in vitro on porcine coronary artery rings. We found that Oly causes endothelial injury with perivascular oedema in the heart and lungs, as well as myocardial haemorrhages in rats. Treatment of porcine coronary artery rings with Oly causes concentration-dependent vasoconstriction and prevents endothelium-mediated relaxation. Using TBC3214 as a selective blocker of the endothelin A receptor, we showed that vasoconstriction induced by Oly was independent of endothelin release and its effects. Verapamil (1 μM) greatly reduced Oly-evoked contractions of porcine coronary artery rings, while lanthanum abolished them completely. These results provide evidence that the contraction of coronary arteries by Oly is due mainly to the increased influx of Ca²⁺ from the extracellular space through voltage-dependent L-type Ca²⁺ channels and cation non-selective channels. Experiments suggest that Oly damages endothelial cells both in vitro and in vivo, and probably exhibits direct contractile effects on coronary smooth muscle cells.     

Efficient oxidation of promutagenic hydroxymethylpyrenes by cDNA-expressed human alcohol dehydrogenase ADH2 and its inhibition by various agents

Alkylated polycyclic aromatic hydrocarbons can be metabolically activated via benzylic hydroxylation and sulphation to electrophilically reactive esters. However, we previously found that the predominant biotransformation route for the hepatocarcinogen 1-hydroxymethylpyrene (1-HMP) in the rat in vivo is the oxidation of the side chain by alcohol dehydrogenases (ADHs) and aldehyde dehydrogenases to the carboxylic acid. Inhibition of this pathway by ethanol (competing ADH substrate) or 4-methylpyrazole (ADH inhibitor) led to a dramatic increase in the 1-HMP-induced DNA adduct formation in rat tissues in the preceding study. In order to elucidate the role of individual ADHs in the metabolism of alkylated polycyclic aromatic hydrocarbons, we expressed the various members of the human ADH family in bacteria. Cytosolic preparations from bacteria expressing ADH2 clearly oxidized hydroxymethylpyrene isomers (1-, 2- and 4-HMP) with the highest rate. This form was purified to near homogeneity to perform detailed kinetic analyses. High catalytic efficiencies (V(max)/K(m)) were observed with HMPs. Thus, this value was 10,000-fold higher for 2-HMP than for the reference substrate, ethanol. The corresponding aldehydes were also efficiently reduced by ADH2. 4-Methylpyrazole inhibited the oxidation of the HMP isomers as well as the reverse reaction. Daidzein, cimetidine and the competing substrate ethanol were further compounds that inhibited the ADH2-mediated oxidative detoxification of 1-HMP.     

Differences in kinetics of donor lymphoid cells in response to G-CSF administration may affect the incidence and severity of acute GvHD in respective HLA-identical sibling recipients

Granulocyte-colony stimulating factor (G-CSF), in addition to myeloid and stem cells, mobilizes a large number of lymphoid cells. We examined which lymphoid populations were mobilized in 21 consecutive donors of peripheral blood stem cells (PBSC) and whether the differences in mobilization could affect the incidence of acute and chronic GvHD in respective HLA-identical recipients. G-CSF administration induced significant increases of donor B (CD3⁻CD19⁺) lymphocytes and slight increases of T (CD3⁺) and cytotoxic (CD16⁺CD56⁺) NK cells. The number of extrathymic cells (CD3⁺ cells with NK markers, or CD7⁺) remained unchanged except for an increase of CD3⁺CD57⁺CD8⁺ cells. Donors of patients without subsequent grade II–IV acute GvHD compared to donors of patients who developed significant acute GvHD were found to have in peripheral blood stable numbers of CD3⁺CD4⁺ cells producing IL2, with a concomitant increased number of CD3⁺CD4^low+CD25⁺ T regulatory cells and decreased NK-mediated cytotoxicity, together with a higher number of suppressive extrathymic CD57⁺CD3⁺ cells in the blood and G-PBMC grafts. Increasing numbers of activated T and NK cells in the blood were associated with the development of chronic GvHD. We suggest that differences in steady-state levels and kinetics of G-CSF induced mobilization of donor lymphoid cells may in addition to other well-known factors affect the incidence of GvHD in HLA-identical recipients. However, owing to the small number of donor-recipient pairs studied, our results must be verified in a larger group of patients. Both authors contributed equally to this study.     

May 2002: 38-year-old man and 69-year-old woman with dural based masses

The May 2002 COM. A 38-year-old man presented with new onset seizures and a 69-year-old woman presented with bilateral headaches and episodes of syncope. Both were found to have extra-axial masses that were contrast-enhancing and thought to be meningiomas. Both had complete resection. Microscopic examination revealed an inflammatory lesion composed of plasma cells, scattered lymphocytes and numerous large histocytic cells, which exhibited emperi polesis and were CD1 a negative, but positive for CD68 and S100. The diagnosis of Destombes-Rosai-Dorfman Disease (DRDD) was rendered. Both cases had good long-term outcome. The differential diagnosis of inflammatory masses in the dura (plasmacytoma, lymphomas, plasma cell fibroma, angiofollicular hyperplasia [Castleman's-disease] and Langerhan's cell histiocytosis) are discussed.     

Purification of a new manganese peroxidase of the white-rot fungus Irpex lacteus, and degradation of polycyclic aromatic hydrocarbons by the enzyme

The white-rot fungus Irpex lacteus has been reported to be an efficient degrader of polycyclic aromatic hydrocarbons, polychlorinated biphenyls and pentachlorophenol. The fungus produces ligninolytic enzymes laccase, lignin peroxidase and manganese peroxidase (MnP), the latter being the major one produced. MnP was purified using anion exchange and size exclusion chromatography. SDS-PAGE showed the purified MnP to be a monomeric protein of 37 kDa (37.5 kDa using MALDI-TOF) with an isoelectric point at 3.55. The pH optimum was relatively broad, from 4.0 to 7.0 with a peak at pH 5.5. Kinetic constants K(m) were 8 microM for H(2)O(2) and 12 or 31 microM for Mn(2+) depending on the substrate. The enzyme did not perform oxidation in the absence of H(2)O(2) or Mn(2+). MnP was active at 5-70 degrees C with an optimum between 50-60 degrees C. At temperatures above 65 degrees C the enzyme rapidly lost activity. Degradation of four representatives of PAHs (phenanthrene, anthracene, fluoranthene, and pyrene) was tested and the enzyme showed the ability to degrade them in vitro. Major degradation products of anthracene were identified. The results confirm the role of MnP in PAH degradation by I. lacteus, including cleavage of the aromatic ring.     

Risk factors for asthma in children in Lódź region

The incidence of asthma and other allergic diseases continues to increase. In addition to genetic factors, environmental influences are thought to play an important role. The aim of this study was to identify factors that influence and drive the atopic march from atopic sensitization to asthma in children from Lód?. METHODS: 800 atopic children, aged 5-18 years, were included to our study. Parents filled in questionnaires and gave interviews about their children's diseases. 405 (43%) children have diagnosis of asthma. RESULTS: A significant association was observed between asthma and male sex, parents' history of atopy, parental highest school grade, maternal smoking during pregnancy, maternal chronic disease (especially chronic renal diseases), maternal allergen-sensitizing diet during breast-feeding, increased exposure to indoor humidity and moulds. Similar effect was seen for episodes of wheeze occurring in the first 3 years of life as followed: wheezing during an airway infection, wheezing not connected with respiratory tract infection, wheezing not related with physical exercise. Child's daycare attendance (nursery school) was associated with decreased risk of asthma.