Cellular distribution of complement receptor type 4 (CR4): expression on human platelets

Neutrophils have been shown to express a receptor for C3dg that is distinct from CR2 and is termed complement receptor type 4 (CR4). In the present study, other peripheral blood cell types were examined by indirect immunofluorescence and flow cytometry for the presence of C3dg binding activity. Specific uptake of C3dg occurred with neutrophils, platelets, and B lymphocytes, but not with eosinophils or T lymphocytes. Monocytes, contained within a mixed cell population of peripheral blood mononuclear cells and platelets, also bound C3dg, whereas purified monocytes did not. Binding of 125I-labeled glutaraldehyde-cross-linked C3dg to platelets was saturable, with an average of 1940 C3dg molecules bound per platelet at saturation (n = 8), ranging in number from 660 to 3930 molecules bound. Activation of platelets with thrombin did not consistently cause an increase in the expression of CR4 sites. 125I-C3dg binding to platelets was competitively inhibited equally well by unlabeled C3dg and iC3b, and approximately fourfold less well by C3b. The addition of platelets to elutriated monocytes generated C3dg binding activity on these cells by the formation of platelet-monocyte complexes. Thus, the CR4 on platelets accounted for the C3dg binding activity initially observed with partially purified monocytes. The adherent property of platelets may enable them to confer on certain other cell types the ability to localize C3dg-coated immune complexes or particles.     

Modifications to United States Environmental Protection Agency Methods 1622 and 1623 for Detection of Cryptosporidium Oocysts and Giardia Cysts in Water

Collaborative and in-house laboratory trials were conducted to evaluate Cryptosporidium oocyst and Giardia cyst recoveries from source and finished-water samples by utilizing the Filta-Max system and U.S. Environmental Protection Agency (EPA) methods 1622 and 1623. Collaborative trials with the Filta-Max system were conducted in accordance with manufacturer protocols for sample collection and processing. The mean oocyst recovery from seeded, filtered tap water was 48.4% ± 11.8%, while the mean cyst recovery was 57.1% ± 10.9%. Recovery percentages from raw source water samples ranged from 19.5 to 54.5% for oocysts and from 46.7 to 70.0% for cysts. When modifications were made in the elution and concentration steps to streamline the Filta-Max procedure, the mean percentages of recovery from filtered tap water were 40.2% ± 16.3% for oocysts and 49.4% ± 12.3% for cysts by the modified procedures, while matrix spike oocyst recovery percentages ranged from 2.1 to 36.5% and cyst recovery percentages ranged from 22.7 to 68.3%. Blinded matrix spike samples were analyzed quarterly as part of voluntary participation in the U.S. EPA protozoan performance evaluation program. A total of 15 blind samples were analyzed by using the Filta-Max system. The mean oocyst recovery percentages was 50.2% ± 13.8%, while the mean cyst recovery percentages was 41.2% ± 9.9%. As part of the quality assurance objectives of methods 1622 and 1623, reagent water samples were seeded with a predetermined number of Cryptosporidium oocysts and Giardia cysts. Mean recovery percentages of 45.4% ± 11.1% and 61.3% ± 3.8% were obtained for Cryptosporidium oocysts and Giardia cysts, respectively. These studies demonstrated that the Filta-Max system meets the acceptance criteria described in U.S. EPA methods 1622 and 1623.     

Regulation of principal cell pH by Na/H exchange in rabbit cortical collecting tubule

Summary Changes in intracellular pH (pH _i ) were measured using the pH indicator, BCECF, in principal cells from split opened cortical collecting tubules (CCTs) derived from rabbits maintained on a normal diet. This monolayer preparation has the advantage of allowing us to visualize the morphological differences in the two major cell types in this nephron segment under transmitted light. The visual identification of the cell types was verified using emission measurements taken from single principal and intercalated cells in the opened tubule which had been exposed to fluorescein isothiocyanate (FITC)-labeled peanut lectin. We confirmed the existence of an amiloride-sensitive Na/H exchange process activated during intracellular acidosis in principal cells. In addition, the exchanger was active under basal conditions and over a wide range of pH _i . Because the exchanger was active under basal conditions we tested the hypothesis that changes in intracellular Na (Na _i ) would alter pH _i in a predictable way. Maneuvers designed to alter Na _i were without significant effects within a 10-min time frame. Specifically, addition of 100 m ouabain to increase Na _i or exposure of the tubules to 10^–5 m amiloride to decrease luminal Na entry and reduce Na _i did not have an effect on pH _i . In some experiments we did observe however, after a 30-min exposure to ouabain, a small decrease in pH _i . These results suggest that Na/H exchange is a major regulator of pH _i in principal cells. However, regulation of Na transport by changes in pH _i in principal cells of rabbit CCT via the activity of a Na/H exchanger do not seem to contribute to the feedback control of Na transport.This work was supported by U.S. Public Health Service grants DK27847 to L.G. Palmer and DK11489 to E.E. Windhager.     

Signaling of Ankle Joint Position by Receptors in Different Muscles

Plots were made of multiunit activity versus ankle joint position for receptors in each of the 12 muscles crossing the cat ankle joint, except peroneus tertius, by recording from populations of afferent fibers in muscle nerves. The discharge was measured 15 or 30 sec after terminating the movements that altered the position of the joint. These recordings were dominated by large-spike activity that would be expected to originate mainly from primary spindle endings. Seven of the 12 muscles also cross other joints. Their responses at a given ankle joint position were so altered by changes in the position of the knee or toe joints that they could not reliably signal the position of the ankle joint. As judged from multiunit recording, receptors in each of the five muscles specific to the ankle joint were influenced by more than one axis of ankle joint displacement.

Single-unit recording from dorsal root filaments was used to determine whether primary or secondary spindle receptors in soleus and tibialis anterior could selectively signal one axis of ankle joint rotation. Individual soleus receptors were tested both on the flexion extension axis and with a combined adduction–eversion movement.

For 38 of the 70 soleus receptors examined (54%), firm adduction–eversion produced a level of activity greater than that caused by 10° of flexion, and for 77% the level of activity was greater than that caused by 5° of flexion. For 168 of the 184 tibialis anterior receptors studied (91%), firm abduction inversion produced a level of activity greater than that caused by 10° of extension. Thus few receptors were found that responded exclusively to one axis of rotation.

One way in which the position of the ankle joint could be specified in the face of multiaxial receptor activity is by examining the receptor discharge from more than one muscle. A suggestion for how the nervous system might do this is given in the discussion.     

Efficiency of Cell-Free and Cell-Associated Virus in Mucosal Transmission of Human Immunodeficiency Virus Type 1 and Simian Immunodeficiency Virus

Effective strategies are needed to block mucosal transmission of human immunodeficiency virus type 1 (HIV-1). Here, we address a crucial question in HIV-1 pathogenesis: whether infected donor mononuclear cells or cell-free virus plays the more important role in initiating mucosal infection by HIV-1. This distinction is critical, as effective strategies for blocking cell-free and cell-associated virus transmission may be different. We describe a novel ex vivo model system that utilizes sealed human colonic mucosa explants and demonstrate in both the ex vivo model and in vivo using the rectal challenge model in rhesus monkeys that HIV-1-infected lymphocytes can transmit infection across the mucosa more efficiently than cell-free virus. These findings may have significant implications for our understanding of the pathogenesis of mucosal transmission of HIV-1 and for the development of strategies to prevent HIV-1 transmission.     

Surveillance of the Sensitivity towards Antiparasitic Bath-Treatments in the Salmon Louse (Lepeophtheirus salmonis)

The evolution of drug resistant parasitic sea lice is of major concern to the salmon farming industry worldwide and challenges sustainable growth of this enterprise. To assess current status and development of L. salmonis sensitivity towards different pesticides used for parasite control in Norwegian salmon farming, a national surveillance programme was implemented in 2013. The programme aims to summarize data on the use of different pesticides applied to control L. salmonis and to test L. salmonis sensitivity to different pesticides in farms along the Norwegian coast. Here we analyse two years of test-data from biological assays designed to detect sensitivity-levels towards the pesticides azamethiphos and deltamethrin, both among the most common pesticides used in bath-treatments of farmed salmon in Norway in later years. The focus of the analysis is on how different variables predict the binomial outcome of the bioassay tests, being whether L. salmonis are immobilized/die or survive pesticide exposure. We found that local kernel densities of bath treatments, along with a spatial geographic index of test-farm locations, were significant predictors of the binomial outcome of the tests. Furthermore, the probability of L. salmonis being immobilized/dead after test-exposure was reduced by odds-ratios of 0.60 (95% CI: 0.42–0.86) for 2014 compared to 2013 and 0.39 (95% CI: 0.36–0.42) for low concentration compared to high concentration exposure. There were also significant but more marginal effects of parasite gender and developmental stage, and a relatively large random effect of test-farm. We conclude that the present data support an association between local intensities of bath treatments along the coast and the outcome of bioassay tests where salmon lice are exposed to azamethiphos or deltamethrin. Furthermore, there is a predictable structure of L. salmonis phenotypes along the coast in the data, characterized by high susceptibility to pesticides in the far north and far south, but low susceptibility in mid Norway. The study emphasizes the need to address local susceptibility to pesticides and the need for restrictive use of pesticides to preserve treatment efficacy.