Correlation of local lightning activity with extra low frequency detector for Schumann Resonance measurements.

A dedicated extremely low frequency (ELF) detector has been constructed and used successfully for Schumann Resonance (SRs) measurements in N.W. Greece. The main objective of this work was to investigate the effect of local lightning activity on the signal of our ELF detector and consequently on the estimated SRs parameters, namely the power, the frequency and the quality factor of each mode. Therefore, several measurements were taken into account for the ELF signal continuously recorded and the lightning intensity in a specific range around the ELF detector. Additionally, a simple filtering technique was used, in order to reject the distorted SRs spectra. The statistical analysis performed showed a positive and statistically significant correlation between the lightning events and the recorded magnetic component of the ELF signal. It was found that local lightnings have a significant impact on the SRs measurements, and it is necessary to be removed from the background signal.

Study of the variations in the Schumann resonances parameters measured in a southern Mediterranean environment.

Schumann resonances (SRs) provide a unique tool for exploring continuous and long-term monitoring of global environmental parameters, although their detection and study is a very complex task. This paper presents diurnal and seasonal variations of principal parameters, namely the resonance frequency, the Q-factor, and the power of SRs, in the North-South (NS) magnetic field component for the first five modes from Extra Low Frequency (ELF) measurements. These were acquired at the observation site located at Doliana, Kalpaki, in the Region of Epirus, North West Greece. These variations are the first ones reported for the principal parameters in the South Eastern Mediterranean area and correspond to the one-year measurement period from 2016 to 2017. The comparison with variations at several observations sites is also discussed. Moreover, correlation with global lighting centers, as well as local lightning activity, and SRs parameters variation is attempted.

Alveolar hydatid cyst (AHC): inflammation-induced reactive gastrointestinal (GL) amyloidosis in AHC-infected mice and chemical characterization of the GL amyloid.

A high incidence of GI amyloidosis has been described in patients with various forms of systemic amyloidosis but its evolution and progression in different subregions of the GI tract are not well documented. These aspects including the chemical nature of GI amyloid were examined in the AHC mouse model of inflammation-associated reactive amyloidosis. C57BL/6 mice were infected intraperitoneally with 250 AHC. Paraffin sections from the stomach and the small and large intestines of AHC mice were stained at different time intervals with Congo red or immunocytochemically with monospecific RAA. The submucosal blood vessels at 1 week postinfection were found to be the first target of amyloid deposition. With time the amyloid deposits extended to the mucosa and the Peyer's patches and immunoreacted with RAA; ileum was the most severely affected region. Amyloid was extracted from the GI tract and purified by size exclusion chromatography using 5 M guanidine-formic acid, pH 3. The purified amyloid was identified by Western blotting using RAA and by partial N-terminal microsequencing up to 10 cycles. The GI amyloid showed homology with murine SAA2, although SAA2 mRNA is not expressed in murine GI tract. These results shows that (a) the GI amyloid is derived, similar to that of splenic/hepatic amyloid, from circulating SAA2 and (b) the GI tract submucosal blood vessels are the first target of AA deposition. The data also suggest that AA-mediated damage to the submucosal blood capillaries may lead to SAA leakage followed by cascading of AA deposition in other layers of the GI tract.

Immunolocalization of serum amyloid A and AA amyloid in lysosomes in murine monocytoid cells: confocal and immunogold electron microscopic studies.

Murine AA amyloid (AA) protein represents the amino-terminal two-third portion of SAA2, one of the isoforms of serum amyloid A. Whether plasma membrane-bound or lysosomal enzymes in activated murine monocytoid cells degrade SAA2 to generate amyloidogenic AA-like peptides is not clearly understood, although AA has been localized in the lysosomes. Here we show, using confocal and immunogold microscopy (IEM), that both SAA and AA localize in lysosomes of activated monocytoid cells from amyloidotic mice. Rabbit anti-mouse AA IgG (RAA) and two monoclonal antibodies against murine lysosome-associated membrane proteins (LAMP-1 and LAMP-2) were used to immunolocalize SAA/AA and lysosomes, respectively. Confocal analysis co-localized both anti-RAA and anti-LAMP-1/LAMP-2 reactivities in the perikaryal organelles which by IEM proved to be electron-dense lysosomes. LAMP-1/LAMP-2-specific gold particles were also localized on lysosomal and perikaryal AA. The results suggest sequestration of SAA into the lysosomes. Since monocytoid cells are not known to phagocytose native amyloid fibrils, our results implicate lysosomes in AA formation.

Ubiquitin profile in inflammatory leukocytes and binding of ubiquitin to murine AA amyloid: immunocytochemical and immunogold electron microscopic studies.

Lysosomes in activated murine monocytoid cells have been implicated in AA amyloid formation. The pathophysiology of this process is not well understood. Previous studies into the nature of the relationship between ubiquitin (UB), possessing intrinsic amyloid enhancing factor (AEF) activity; serum amyloid A (SAA), the precursor protein of AA amyloid; and activated monocytoid cells have indicated a temporal and spatial relationship between these proteins and tissue AA amyloid deposits. To extend these findings, we have examined murine peritoneal leukocytes and splenic tissues during the early amyloid deposition phase by immunocytochemical and immunogold electron microscopic methods using monospecific anti-ubiquitin and anti-mouse AA amyloid antibodies. We show here enrichment of endosome-lysosome-like (EL) vesicles in the activated monocytoid cells with UB and SAA, and the presence of UB-bound AA amyloid fibrils in the EL vesicles, perikarya, and interstitial spaces. The importance of these findings is emphasized by the fact that activated monocytoid cells, containing UB in the EL vesicles, sequester and eventually localize SAA in their EL vesicles, and that UB binds to the EL-contained AA amyloid fibrils. These findings may also have functional consequences for studies on the role of EL and UB in amyloidogenesis.

Ubiquitin: its potential significance in murine AA amyloidogenesis.

Amyloid enhancing factor (AEF), which has recently been shown to have identity with ubiquitin (Ub), is believed to play a causative role in experimentally induced AA amyloidosis in mice. We have examined the profile of Ub in activated leukocytes and splenic reticulo-endothelial (RE) cells and its relationship with serum amyloid A protein (SAA) and AA amyloid deposits in an alveolar hydatid cyst (AHC)-infected mouse model of AA amyloidosis. Two monospecific antibodies, anti-ubiquitin (RABU) and anti-mouse AA amyloid, were used as immunological probes to localize Ub, SAA, and AA amyloid. In response to AHC infection, the dull and diffuse Ub immunoreactivity in normal mouse leukocytes and RE cells promptly changed to a discrete granular pattern suggesting an increase in the intracellular concentration of Ub and the formation of Ub-protein conjugates. This corresponded to an elevation in SAA levels, SAA uptake by RABU-positive phagocytic cells, co-localization of Ub-SAA immunoreactive splenocytes in the perifollicular areas, and deposition of Ub-bound AA amyloid in the splenic and hepatic tissues. These results suggest that Ub-loaded monocytoid cells may play an important role in the physiological processing of the sequestered SAA into AA amyloid. Aspects of AA amyloidogenesis are discussed in relation to other experimental models in which stress-induced Ub-protein conjugate formation and its transport to lysosomal vesicles have been studied.

Ubiquitin profile and amyloid enhancing factor activity in Alzheimer and 'normal' human brain extracts.

Tris-HCl or Laemmli sample buffer extracted frontal lobe and hippocampal samples from normal aged and Alzheimer's disease (AD) subjects were used to determine total ubiquitin (Ub), distribution of monomeric Ub and Ub-protein conjugates and amyloid enhancing factor (AEF) activity using the dot-blot, Western blot and mouse AEF bioassay techniques, respectively. The AD samples, as compared to the normals, demonstrated a 1.7-fold increase in total Ub, elevated levels of Ub-protein conjugates and an appreciably enhanced AEF activity. Many of the hippocampal Ub-protein conjugates were found to be soluble only in the Laemmli sample buffer. The possible roles of elevated Ub levels and of the association of AEF activity with Ub are discussed in regard to pathogenesis of brain amyloidosis.

Amyloid enhancing factor activity is associated with ubiquitin.

Crude amyloid enhancing factor (AEF) drastically reduces the pre-amyloid phase on passive transfer and induces amyloid deposition in the recipient mice in 48-120 h. We attempted to purify AEF from murine amyloidotic liver and spleen extracts by using gel filtration, preparative sodium dodecyl sulphate-polyacrylamide gel electrophoresis and ion exchange chromatography and isolated a 5.5 kDa peptide. In the mouse bioassay, this peptide induced accelerated splenic AA deposition in a dose-dependent manner. Based on structural, electrophoretic and immunochemical criteria the peptide was identified as ubiquitin. A polyclonal rabbit anti-bovine ubiquitin IgG antibody (RABU) abolished the in vivo AEF activity of crude murine AEF in a dose-dependent manner. Monomeric ubiquitin and its large molecular weight adducts were isolated from crude AEF using cyanogen bromide-activated sepharose conjugated to RABU and size exclusion chromatography methods. These were assayed and were found to possess AEF activity. Furthermore, increased levels of ubiquitin, a phenomenon similar to that of AEF, were detected by immunocytochemistry in mouse peritoneal leucocytes prior to and during amyloid deposition. Since AEF shares a number of biological and functional properties with ubiquitin, we suggest a possible role of ubiquitin as an AEF, and that serum amyloid protein A and ubiquitin, the two reactants generated during inflammatory stress conditions, may converge to induce AA amyloid deposition.

Binding of ubiquitin to experimentally induced murine AA amyloid.

Amyloid enhancing factor (AEF) activity has recently been demonstrated in ubiquitin purified from amyloidotic murine tissues and Alzheimer brain extract. Since AEF is known to bind to amyloid fibrils and 'fibril-AEF' on passive transfer induces accelerated amyloidogenesis in the recipient animals, it was of interest to investigate whether ubiquitin binds to amyloid. Immunohistological studies were carried out on liver sections from amyloidotic mice. Biotin-strepavidin-peroxidase methods using monospecific rabbit anti-mouse AA amyloid IgG (RAAG) and rabbit anti-bovine ubiquitin IgG (RABU) antibodies were employed to immunostain the amyloid and ubiquitin deposits, respectively. RABU-treated liver sections were counterstained with thioflavine S. RAAG reacted strongly with the amyloid, indicating that it is AA type, and RABU-positive immunodeposits were found bound to the thioflavine-S-positive AA deposits. Treatment of the liver sections with 0.1 M sodium acetate containing 0.5 M NaCl, pH 4, for 2-3 h at 37 degrees C nearly completely desorbed the AA amyloid-bound ubiquitin. Since ubiquitin demonstrates AEF activity in vivo and binds non-covalently to AA amyloid, we suggest that ubiquitin may indeed be 'fibril-AEF' and may play a crucial role in the pathogenesis of amyloidosis. To our knowledge, this is the first time that ubiquitin bound to extracellularly deposited amyloid has been demonstrated.

Alzheimer's disease brain-derived ubiquitin has amyloid-enhancing factor activity: behavior of ubiquitin during accelerated amyloidogenesis.

Amyloid-enhancing factor (AEF) is believed to be a crucial common pathogenetic link in diverse forms of human amyloidosis. Passive transfer of crude AEF is known to trigger accelerated splenic amyloid deposition in mice. We have recently identified AEF activity in ubiquitin isolated from murine amyloidotic tissues. Using similar techniques we have purified ubiquitin, from crude Alzheimer's disease (AD) brain extracts, to apparent homogeneity. Based on the partial amino acid sequence homology, immunochemical and pathophysiological criteria, the approximately 5.5-kDa AD-derived protein was identified as ubiquitin (AD-ubiquitin) with AEF activity. Ten to twenty micrograms of this protein per mouse, with or without CaCl2, in conjunction with four subcutaneous injections of 0.5 ml of 1% aqueous AgNO3, induced accelerated splenic amyloid deposition. By immunohistochemistry, using anti-mouse AA amyloid antibody, the AD-ubiquitin-induced amyloid was identified as AA type. With anti-bovine ubiquitin antibody, using similar spleen sections as above, ubiquitin was found to co-deposit with AA amyloid in the splenic perifollicular areas. These results strongly suggest that ubiquitin may be involved in the pathogenesis of amyloidosis.