Circulating inflammatory mediators during start of fever in differential diagnosis of gram-negative and gram-positive infections in leukopenic rats

Gram-negative and gram-positive infections have been considered the most important causes of morbidity and mortality in patients with leukopenia following chemotherapy. However, discrimination between bacterial infections and harmless fever episodes is difficult. Because classical inflammatory signs of infection are often absent and fever is frequently the only sign of infection, the aim of this study was to assess the significance of serum interleukin-6 (IL-6), IL-10, macrophage inflammatory protein-2 (MIP-2), procalcitonin (PCT), and C-reactive protein (CRP) patterns in identifying bacterial infections during start of fever in normal and cyclophosphamide-treated (leukopenic) rats following an injection of lipopolysaccharide (LPS) or muramyl dipeptide (MDP) as a model for gram-negative and gram-positive bacterial infections. We found that, compared to normal rats, immunosuppressed animals exhibited significantly higher fevers and lesser production of all mediators, except IL-6, after toxin challenge. Moreover, compared to rats that received MDP, both groups of animals that received an equivalent dose of LPS showed significantly higher fevers and greater increase in serum cytokine levels. Furthermore, in contrast to those in immunocompetent rats, serum levels of IL-6 and MIP-2 were not significantly changed in leukopenic animals after MDP injection. Other serum markers such as PCT and CRP failed to discriminate between bacterial stimuli in both groups of animals. These results suggest that the use of the analyzed serum markers at an early stage of fever could give useful information for the clinician for excluding gram-negative from gram-positive infections.     

Blinded, Externally Controlled Multicenter Evaluation of Light Microscopy and PCR for Detection of Microsporidia in Stool Specimens

The quality parameters for the detection of microsporidia in identical sets of 50 stool samples were determined for six laboratories where technicians used light microscopy and for six laboratories where technicians used PCR. The average overall sensitivities were 67% (89% for patient samples only) for the PCR laboratories and 54% (80% for patient samples only) for the light microscopy laboratories. Specificities were 98 and 95%, respectively. Differences in results were most apparent between the individual laboratories rather than between the two major methods used.     

Epidemiology of Cyclospora cayetanensis and other intestinal parasites in a community in Haiti

We conducted an exploratory investigation in a community in Haiti to determine the prevalence of Cyclospora cayetanensis infection and to identify potential risk factors for C. cayetanensis infection. In 2001, two cross-sectional stool surveys and a nested case-control study were conducted. In 2002, a follow-up cross-sectional stool survey was conducted among children < or =10 years of age. Stool specimens from study participants and water samples from their wells were examined for Cyclospora and other intestinal parasites. In stools, the prevalence of infection with Cyclospora in persons of all ages decreased from 12% (20 of 167 persons) in February 2001 to 1.1% (4 of 352 persons) in April 2001, a 90.8% decrease. For children < or =10 years of age, the prevalence rates were 22.5% (16 of 71 children) in February 2001, 3.0% (4 of 135 children) in April 2001, and 2.5% (2 of 81 children) in January 2002. Use of the water from the artesian well in the northern region of the community versus the one in the south was the only risk factor associated with Cyclospora infection in multivariate analyses (odds ratio, 18.5; 95% confidence interval, 2.4 to 143.1). The water sample from one of the nine wells or water sources tested (one sample per source) in January 2001, shortly before the investigation began, was positive for Cyclospora by UV fluorescence microscopy and PCR. None of the water samples from the 46 wells or water sources tested during the investigation (one sample per source per testing period, including the artesian wells) were positive for Cyclospora. Further studies are needed to assess the role of water as a possible risk factor for Cyclospora infection in Haiti and other developing countries.     

Regulation of human basophil adhesion to endothelium under flow conditions: Different very late antigen 4 regulation on umbilical cord blood-derived and peripheral blood basophils

BACKGROUND: Although soluble mediators released by basophils in tissue sites contribute to the chronic injury that occurs in hypersensitivity diseases, only limited information is available about how circulating basophils are recruited to tissues. In particular, the interaction of basophils with endothelium under conditions that mimic physiologic flow has not been explored. OBJECTIVE: We sought to identify adhesion molecules regulating the attachment of human basophils to IL-4-activated human umbilical vein endothelial cells (HUVECs) under flow conditions. METHODS: A parallel-plate flow chamber and blocking mAbs were used to define the adhesion molecules involved in the interactions of peripheral blood basophils (PBBs) and cord blood-derived basophils (CBDBs) with IL-4-activated HUVECs and with Chinese hamster ovary (CHO) cell transfectants expressing specific adhesion molecules. A fluorescent ligand specific for very late antigen 4 (VLA-4) was used to directly examine the VLA-4 affinity state of basophils. RESULTS: Flowing PBBs and CBDBs attached to activated HUVECs and to CHO cells expressing P- or E-selectin. However, only CBDBs attached to vascular cell adhesion molecule 1 (VCAM-1)-transfected CHO cells under flow conditions. The attachment of CBDBs to CHO cells was blocked by mAbs directed against E-selectin, P-selectin, and VCAM-1, whereas attachment of PBBs was blocked by E-selectin and P-selectin mAbs. Activating VLA-4 with Mn(2+) on PBBs resulted in adhesion to the VCAM-1-transfected CHO cells, indicating that VLA-4 activity on PBBs can be regulated, at least in part, through affinity changes. The Mn(2+)-induced upregulation of basophil VLA-4 affinity was demonstrated directly by using a fluorescent ligand for VLA-4 and flow cytometry. CONCLUSIONS: The interaction of human CBDBs and PBBs with endothelium under flow conditions is mediated in part by both P- and E-selectin. VLA-4 additionally contributes to the adhesion of flowing CBDBs. However, the affinity of VLA-4 is too low to support the adhesion under flow conditions of unstimulated PBBs.     

Do semantic categories activate distinct cortical regions? Evidence for a distributed neural semantic system

A key issue in cognitive neuroscience concerns the neural representation of conceptual knowledge. Currently, debate focuses around the issue of whether there are neural regions specialised for the processing of specific semantic attributes or categories, or whether concepts are represented in an undifferentiated neural system. Neuropsychological studies of patients with selective semantic deficits and previous neuroimaging studies do not unequivocally support either account. We carried out a PET study to determine whether there is any regional specialisation for the processing of concepts from different semantic categories using picture stimuli and a semantic categorisation task. We found robust activation of a large semantic network extending from left inferior frontal cortex into the inferior temporal lobe and including occipital cortex and the fusiform gyrus. The only category effect that we found was additional activation for animals in the right occipital cortex, which we interpret as being due to the extra visual processing demands required in order to differentiate one animal from another. We also carried out analyses in specific cortical regions that have been claimed to be preferentially activated for various categories, but found no evidence of any differential activation as a function of category. We interpret these data within the framework of cognitive accounts in which conceptual knowledge is represented within a nondifferentiated distributed system.     

Functional proteomics mapping of a human signaling pathway

Access to the human genome facilitates extensive functional proteomics studies. Here, we present an integrated approach combining large-scale protein interaction mapping, exploration of the interaction network, and cellular functional assays performed on newly identified proteins involved in a human signaling pathway. As a proof of principle, we studied the Smad signaling system, which is regulated by members of the transforming growth factor beta (TGFbeta) superfamily. We used two-hybrid screening to map Smad signaling protein-protein interactions and to establish a network of 755 interactions, involving 591 proteins, 179 of which were poorly or not annotated. The exploration of such complex interaction databases is improved by the use of PIMRider, a dedicated navigation tool accessible through the Web. The biological meaning of this network is illustrated by the presence of 18 known Smad-associated proteins. Functional assays performed in mammalian cells including siRNA knock-down experiments identified eight novel proteins involved in Smad signaling, thus validating this integrated functional proteomics approach.     

Leishmania major-Like Antigen for Specific and Sensitive Serodiagnosis of Human and Canine Visceral Leishmaniasis

An antigen (LMS) prepared from Leishmania major-like promastigotes was used in an enzyme-linked immunosorbent assay (ELISA) for the diagnosis of human and dog visceral leishmaniasis. The results were compared with those from the indirect immunofluorescent antibody test (IFAT). A total of 1,822 canine sera were tested, including sera from dogs with visceral leishmaniasis, transmissible venereal tumors, ehrlichiosis, rickettsiosis, or Chagas' disease and sera from healthy dogs. The antigen was also tested with 227 samples of human sera, including sera from patients with visceral, cutaneous, or diffuse cutaneous leishmaniasis and from noninfected individuals, as well as sera from patients with Chagas' disease, toxoplasmosis, rickettsiosis, hepatitis B, schistosomiasis, ascaridiasis, malaria, rheumatoid factor, leprosy and rheumatoid factor, tuberculosis, or leprosy. All dogs and all human patients had a clinical and/or serological and/or parasitological diagnosis. For detecting antibodies in sera from dogs with leishmaniasis, the antigen showed a sensitivity of 98%, specificity of 95%, and concordance of 93% and when used for detecting antibodies in human sera presented a sensitivity of 92%, specificity of 100%, and concordance of 92%. Comparison between ELISA and IFAT demonstrated that ELISA using the LMS antigen yielded more reliable results than IFAT. The LMS antigen displayed no cross-reactivity with sera from patients or dogs that had any of the other diseases tested.     

Deposition and passage of transthyretin through the blood-nerve barrier in recipients of familial amyloid polyneuropathy livers

Familial amyloid polyneuropathy (FAP) is characterized by deposition of mutated transthyretin (TTR) in the peripheral nervous system. Prior to amyloid fibrils, nonfibrillar TTR aggregates are deposited inducing oxidative stress with increased nitration (3-NT). As the major source of TTR is the liver, liver transplantation (LT) is used to halt FAP. Given the shortage of liver donors, domino LT (DLT) using FAP livers is performed. The correlation between TTR deposition in the skin and nerve was tested in biopsies from normal individuals, asymptomatic carriers (FAP 0) and FAP patients; in FAP 0, nonfibrillar TTR was observed both in the skin and nerve in the same individuals; in patients, amyloid was detected in both tissues. The occurrence of amyloidosis in recipients of FAP livers was evaluated 1-7 years after DLT: TTR deposition occurred in the skin 3 years after transplantation either as amyloid or aggregates; in one of the recipients, fibrillar TTR was present in the epineurium 6 years after DLT. Deposits were scarce and 3-NT immunostaining was irrelevant. Nerve biopsies from DLT recipients had no FAP-related neuropathy. Our findings suggest that TTR amyloid formation occurs faster than predicted and that TTR of liver origin can cross the blood-nerve barrier. Recipients of FAP livers should be under surveillance for TTR deposition and tissue damage.