THE ELECTROPHORETIC MOBILITY OF RABBIT ERYTHROCYTES AND GHOSTS

Measurements of the electrical mobility of washed rabbit red cells and of ghosts produced by hypotonic solutions, freezing-and-thawing, chloroform, and saponin were made in the Abramson horizontal microelectrophoresis cell. These different forms of lysis, which corresponds to a variety of degrees of injury to the red cell, are unaccompanied by any change in electrical mobility. These observations are discussed from the standpoint of the possible structure of the cell membrane and the action of lysins upon it.     

Lactate inhibits Ca2+-activated Ca2+-channel activity from skeletal muscle sarcoplasmic reticulum

Favero, Terence G., Anthony C. Zable, David Colter, andJonathan J. Abramson. Lactate inhibits Ca²⁺-activatedCa²⁺-channel activity from skeletal muscle sarcoplasmicreticulum. J. Appl. Physiol. 82(2): 447-452, 1997.Sarcoplasmic reticulum (SR) Ca²⁺-release channelfunction is modified by ligands that are generated during about ofexercise. We have examined the effects of lactate on Ca²⁺-and caffeine-stimulated Ca²⁺ release,[³H]ryanodine binding, and singleCa²⁺-release channel activity of SR isolated from rabbitwhite skeletal muscle. Lactate, at concentrations from 10 to 30 mM,inhibited Ca²⁺- and caffeine-stimulated[³H]ryanodine binding to and inhibited Ca²⁺-and caffeine-stimulated Ca²⁺ release from SR vesicles.Lactate also inhibited caffeine activation of single-channel activityin bilayer reconstitution experiments. These findings suggest thatintense muscle activity, which generates high concentrations oflactate, will disrupt excitation-contraction coupling. This may lead todecreases in Ca²⁺ transients promoting a decline in tensiondevelopment and contribute to muscle fatigue.

     

Decreased Langerhans Cell Responses to IL-36γ: Altered Innate Immunity in Patients with Recurrent Respiratory Papillomatosis

Recurrent respiratory papillomatosis (RRP) is a rare, chronic disease caused by human papillomaviruses (HPVs) types 6 and 11 that is characterized by the polarization of adaptive immune responses that support persistent HPV infection. Respiratory papillomas express elevated mRNA levels of IL-36γ, a proinflammatory cytokine in comparison to autologous clinically normal laryngeal tissues; however there is no evidence of inflammation in these lesions. Consistent with this, respiratory papillomas do not contain T_H1-like CD4⁺ T-cells or cytotoxic CD8⁺ T-cells, but instead contain a predominance of T_H2-like and T regulatory cells (Tregs). In addition, papillomas also are infiltrated with immature Langerhans cells (iLCs). In this study, we show that papilloma cells express IL-36γ protein, and that human keratinocytes transduced with HPV11 have reduced IL-36γ secretion. We now provide the first evidence that peripheral blood-derived iLCs respond to IL-36γ by expressing inflammatory cytokines and chemokines. When stimulated with IL-36γ, iLCs from patients with RRP had lower expression levels of the T_H2-like chemokine CCL-20 as compared with controls. Patients’ iLCs also had decreased steady state levels of CCL-1, which is a proinflammatory chemokine. Moreover, CCL-1 levels in iLCs inversely correlated with the severity of RRP. The combined decrease of T_H1- and a T_H2-like chemokines by iLCs from patients could have consequences in the priming of IFN-γ expression by CD8⁺ T-cells. Taken together, our results suggest that, in RRP, there is a defect in the proinflammatory innate immune responses made by iLCs in response to IL-36γ. The consequence of this defect may lead to persistent HPV infection by failing to support an effective HPV-specific, T_H1-like and/or T_c1-like adaptive response, thus resulting in the predominant T_H2-like and/or Treg micromilieu present in papillomas.     

Differential Expression of Alpha 4 Integrins on Effector Memory T Helper Cells during Bordetella Infections. Delayed Responses in Bordetella pertussis

Bordetella pertussis (B. pertussis) is the causative agent of whooping cough, a respiratory disease that is reemerging worldwide. Mechanisms of selective lymphocyte trafficking to the airways are likely to be critical in the immune response to this pathogen. We compared murine infection by B. pertussis, B. parapertussis, and a pertussis toxin-deleted B. pertussis mutant (BpΔPTX) to test the hypothesis that effector memory T-helper cells (emTh) display an altered pattern of trafficking receptor expression in B. pertussis infection due to a defect in imprinting. Increased cell recruitment to the lungs at 5 days post infection (p.i.) with B. parapertussis, and to a lesser extent with BpΔPTX, coincided with an increased frequency of circulating emTh cells expressing the mucosal-associated trafficking receptors α4β7 and α4β1 while a reduced population of these cells was observed in B. pertussis infection. These cells were highly evident in the blood and lungs in B. pertussis infection only at 25 days p.i. when B. parapertussis and BpΔPTX infections were resolved. Although at 5 days p.i., an equally high percentage of lung dendritic cells (DCs) from all infections expressed maturation markers, this expression persisted only in B. pertussis infection at 25 days p.i. Furthermore, at 5 days p.i with B. pertussis, lung DCs migration to draining lymph nodes may be compromised as evidenced by decreased frequency of CCR7⁺ DCs, inhibited CCR7-mediated in vitro migration, and fewer DCs in lung draining lymph nodes. Lastly, a reduced frequency of allogeneic CD4⁺ cells expressing α4β1 was detected following co-culture with lung DCs from B. pertussis-infected mice, suggesting a defect in DC imprinting in comparison to the other infection groups. The findings in this study suggest that B. pertussis may interfere with imprinting of lung-associated trafficking receptors on T lymphocytes leading to extended survival in the host and a prolonged course of disease.     

Diffusion and home range parameters from rodent population measurements in Panama

Simple random walk considerations are used to interpret rodent population data collected in Hantavirus-related investigations in Panama regarding the short-tailed cane mouse, Zygodontomys brevicauda. The diffusion constant of mice is evaluated to be of the order of (and larger than) 200 meters squared per day. The investigation also shows that the rodent mean square displacement saturates in time, indicating the existence of a spatial scale which could, in principle, be the home range of the rodents. This home range is concluded to be of the order of 70 meters. Theoretical analysis is provided for interpreting animal movement data in terms of an interplay of the home ranges, the diffusion constant, and the size of the grid used to monitor the movement. The study gives impetus to a substantial modification of existing theory of the spread of the Hantavirus epidemic which has been based on simple diffusive motion of the rodents, and additionally emphasizes the importance for developing more accurate techniques for the measurement of rodent movement.     

An indirect enzyme immunoassay for the mycotoxin citrinin.

An indirect competitive enzyme immunoassay using rabbit antisera could detect citrinin in buffer solutions at 1 to 13 ng/ml (0.05 to 0.65 ng per assay). Cross-reactivity with austdiol, alternariol, ochratoxin A, and deoxynivalenol was < 0.1% relative to citrinin. Recovery of citrinin added to wheat flour at 200 to 2,000 ng/g was 89 to 104%, with a coefficient of variation of 6.9 to 13%.     

Electrophoretic mobility of sarcoplasmic reticulum vesicles is determined by amino acids of A + P + N domains of Ca-ATPase

Establishing the origin of electrophoretic mobility of sarcoplasmic reticulum (SR) vesicles is the primary goal of this work. It was found that the electrophoretic mobility originates from ionizable amino acids of cytoplasmic domains of the Ca²⁺-ATPase, the calcium pump of SR. The mobility was measured at pH 4.0, 4.7, 5.0, 6.0, 7.5, and 9.0 in the region of ionic strength from 0.05 to 0.2 M. Mobility measurements were supplemented by studies of SR vesicles by photoelectron microscopy. The median diameter of SR vesicles was 260 nm. Ca²⁺-ATPases were not resolved. The mobility data were standardized by interpolation to a reference ionic strength of 0.1 M. The mobility of the SR vesicles is determined by the charge of the Ca²⁺-ATPase. It is due to the ionizable amino acids selected from the amino acid sequence of SERCA1a Ca²⁺-ATPase. The pH dependence of charge residing in various domains of Ca²⁺-ATPase was computed using pKa values in free water. The charge correlated with measured mobility. It was shown that a linear relationship exists between the mobility of the SR vesicles, μ, and the total computed charge, Q, on three cytoplasmic domains of Ca²⁺-ATPase: A, P, and N. It is given by μ = α + β Q where the fitted values β = (0.043 ± 0.002) × 10⁻⁸ m² V⁻¹ s⁻¹ e⁻¹ and α = (0.16 ± 0.02) × 10⁻⁸ m² V⁻¹ s⁻¹. Since β and α values do not change from pH 4 to pH 9, one concludes that the hydrodynamic friction of the cytoplasmic domains of SR is independent of their charge.     

Genetic variation and phylogeography of the bank vole (<Emphasis Type="Italic">Clethrionomys glareolus</Emphasis>, Arvicolinae,Rodentia) in Russia with special reference to the introgression of the mtDNA of a closely related species,red-backed vole (<Emphasis Type="Italic">Cl. rutilus</Emphasis>)

Totally, 294 bank voles (Clethrionomys glareolus) and 18 red-backed voles (Cl. rutilus) from 62 sites of European Russia were studied. Incomplete sequences (967 bp) of the mitochondrial cytochrome b gene were determined for 93 Cl. glareolus individuals from 56 sites and 18 Cl. rutilus individuals from the same habitats. Analysis of the cytochrome b gene variation has demonstrated that practically the entire European part of Russia, Ural, and a considerable part of Western Europe are inhabited by bank voles of the same phylogroup, displaying an extremely low genetic differentiation. Our data suggest that Cl. glareolus very rapidly colonized over the presently occupied territory in the post-Pleistocene period from no more than two (central European and western European) refugia from ancestral populations with a small effective size. PCR typing of the mitochondrial cytochrome b gene allowed us to assess the scale of mtDNA introgression from a closely related species, Cl. rutilus, and to outline the geographical zone of this introgression. Comparison with the red-backed vole haplotypes in the habitats shared by both species favors the hypothesis of an ancient hybridization event (mid-Holocene) and a subsequent introgression. These results suggest that the hybridization took place in the southern and middle Pre-Ural region.     

Metabolites of ochratoxins in rat urine and in a culture of Aspergillus ochraceus.

We studied the metabolic profile of ochratoxin A (OA) in rats and in a culture of OA-producing Aspergillus ochraceus. Ochratoxin alpha (O alpha), ochratoxin beta (O beta), 4-R-hydroxyochratoxin A (4-R-OH OA), 4-R-hydroxyochratoxin B (4-R-OH OB), and 10-hydroxyochratoxin A (10-OH OA) were isolated from a culture of A. ochraceus and structurally characterized by 1H nuclear magnetic resonance spectroscopy, mass spectrometry and high-pressure liquid chromatography. 4-R-OH OA and O alpha were consistently produced and were the dominant biotransformed metabolites in the fungal culture and in rats treated with OA and ochratoxin C (OC), while the formation of 10-OH OA was conditional in the fungal system. Green fluorescent biomacromolecules were isolated by detergent extraction of the fungal culture followed by cold-acetone precipitation and gel filtration. Acid hydrolysis of the fluorescent macromolecules resulted in the release of several ochratoxins, including O alpha (80%), OA (2%), and OC (5%), and other unidentified fluorescent compounds but not OB and O beta. Cross-reactivity studies of the natural macromolecule conjugates of OA with anti-OA polyclonal antibodies indicated that they were covalently linked to the macromolecules via a group other than the carboxyl group. These studies demonstrated that a fungus can produce some of the same metabolites of OA as the rat and that O alpha, OA, and OC may be covalently linked to fungal macromolecules.