The role of interleukin 12 in the development of atherosclerosis in ApoE-deficient mice.
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The cytokine profile of atherosclerotic aortas from apoE-deficient mice was assessed by reverse transcriptase-polymerase chain reaction. The results clearly showed that the expression of mRNA for IL-12p40 was evident in aortas from 3-month-old apoE-deficient mice. The mRNA for IL-10 was detected in aorta from these mice at the age of 6 months, indicating that expression of IL-12 is earlier than that of IL-10 in these animals. Concurrent with IL-12p40, the mRNA for the T-cell cytokine IFN-gamma, but not IL-4, was detected in aortas of mice at young and old ages. Both in situ hybridization and immunostaining further demonstrated the localization of IL-12 in macrophages of atherosclerotic lesions. Immunohistochemistry also demonstrated the expression of costimulatory molecules B7-1 and B7-2 in macrophages, suggesting that activation of T lymphocytes by macrophages may occur via surface antigens in lesions. When the immunoglobulin isotype of the antioxidized LDL antibodies in sera of apoE-deficient mice was determined, it revealed that both IgM and IgG were present. Furthermore, IgG2a is predominant and comprises approximately 50% of the antioxidized LDL IgG in sera from young mice (3 months), but decreased to lower levels (35%) in older mice (6 months). Daily administration of IL-12 led to an increase in serum levels of antioxidized LDL antibodies and accelerated atherosclerosis in young apoE-deficient mice compared with control mice injected with PBS alone. Taken together, these data suggest that IL-12 plays an active role in regulating the immune response during the early phase of atherosclerosis in apoE-deficient mice. (+info)
Detection of Bartonella henselae DNA by two different PCR assays and determination of the genotypes of strains involved in histologically defined cat scratch disease.
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Cat scratch disease (CSD) is a common cause of subacute regional lymphadenopathy, not only in children but also in adults. Serological and molecular studies demonstrated that Bartonella henselae is the etiologic agent in most cases of CSD. Amplification of B. henselae DNA in affected tissue and detection of antibodies to B. henselae are the two mainstays in the laboratory diagnosis of CSD. We designed a retrospective study and investigated formalin-fixed, paraffin-embedded lymph nodes from 60 patients (25 female, 35 male) with histologically suspected CSD by PCR amplification. The sensitivities of two different PCR assays were compared. The first primer pair amplified a 296-bp fragment of the 16S rRNA gene in 36 of the 60 samples, corresponding to a sensitivity of 60%. The second primer pair amplified a 414-bp fragment of the htrA gene in 26 of the 60 lymph nodes, corresponding to a sensitivity of 43.3%. Bartonella DNA could be detected in a total of 39 (65%) of the 60 lymph nodes investigated. However, histopathologic findings are typical but not specific for CSD and cannot be considered as a "gold standard" for diagnosis of CSD. The sensitivity of the PCR assays increased from 65 to 87% if two criteria (histology and serology) were used in combination for diagnosis of CSD. Two genotypes (I and II) of B. henselae are described as being involved in CSD. Genotype I was found in 23 (59%) and genotype II was found in 9 (23%) of the 39 PCR-positive lymph nodes. Seven (18%) lymph nodes were negative in both type-specific PCR assays. Thirty (50%) of our 60 patients were younger than 20 years old (15 were younger than 10 years), 20 (33%) were between 21 and 40 years old, and 10 (17%) patients were between 41 and 84 years old. Our data suggest that detection of Bartonella DNA in patients' samples might confirm the histologically suspected diagnosis of CSD. (+info)
Coronary angioplasty induces rise in Chlamydia pneumoniae-specific antibodies.
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Chlamydia pneumoniae is frequently found in atherosclerotic lesions, and high titers of specific antibodies are associated with increased risk for acute myocardial infarction. However, a causative relation has not been established yet. We performed a prospective study of 93 patients undergoing percutaneous transluminal coronary angioplasty (PTCA) to investigate whether angioplasty influences Chlamydia-specific antibody titers and whether there is an association with restenosis. Blood samples were obtained before and 1 and 6 months after angioplasty. Antibodies against chlamydial lipopolysaccharide and against purified C. pneumoniae elementary bodies were measured by enzyme-linked immunosorbent assay (ELISA). After angioplasty, the prevalence of antibodies to lipopolysaccharide rose from 20 to 26% for immunoglobulin A (IgA), from 53 to 64% for IgG, and from 2 to 7% for IgM (P = 0.021, 0.004, and 0.046, respectively). There was a rapid increase of mean antibody titers of all antibody classes within 1 month of PTCA. During the following 5 months, antibody titers decreased slightly but were still higher than baseline values. Results of the C. pneumoniae-specific ELISA were essentially the same. The rise of anti-Chlamydia antibodies was not caused by unspecific reactivation of the immune system, as levels of antibodies against cytomegalovirus did not change. Neither seropositivity nor antibody titers were related to restenosis. However, increases in mean IgA and IgM titers were restricted to patients who had suffered from myocardial infarction earlier in their lives. In conclusion, we show that PTCA induces a stimulation of the humoral immune response against C. pneumoniae. These data support the idea that plaque disruption during angioplasty might make hidden chlamydial antigens accessible to the immune system. (+info)
Indirect enzyme-linked immunosorbent assay for detection of immunoglobulin G reactive with a recombinant protein expressed from the gene encoding the 116-kilodalton protein of Mycoplasma pneumoniae.
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Serology remains the method of choice for laboratory diagnosis of Mycoplasma pneumoniae infection. Currently available serological tests employ complex cellular fractions of M. pneumoniae as antigen. To improve the specificity of M. pneumoniae diagnosis, a recombinant protein was assessed as a serodiagnostic reagent. A panel of recombinant proteins were expressed from a cloned M. pneumoniae gene that encodes a 116-kDa surface protein antigen. The recombinant proteins were assessed for reactivity with patient sera and the most antigenic was further assessed for its serodiagnostic potential by indirect enzyme-linked immunosorbent assay (ELISA). The ELISA based on the recombinant protein was equivalent in sensitivity to the commercial test (Serodia Myco II; Fujirebio Inc.) to which it was compared. Southern and Western blotting data suggested that the recombinant protein derived from the 116-kDa protein of M. pneumoniae could provide a species-specific diagnostic tool, although further assessment is required. (+info)
Rapid and sensitive detection of immunoglobulin M (IgM) and IgG antibodies against canine distemper virus by a new recombinant nucleocapsid protein-based enzyme-linked immunosorbent assay.
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Canine distemper morbillivirus (CDV) infection causes a frequently fatal systemic disease in a broad range of carnivore species, including domestic dogs. In CDV infection, classical serology provides data of diagnostic and prognostic values (kinetics of seroconversion) and is also used to predict the optimal vaccination age of pups. Routine CDV serology is still based on time- and cost-intensive virus neutralization assays (V-NA). Here, we describe a new capture-sandwich enzyme-linked immunosorbent assay (ELISA) that uses recombinant baculovirus-expressed nucleocapsid (N) protein of a recent CDV wild-type isolate (2544/Han95) for the detection of CDV-specific antibodies in canine sera. Recombinant antigen was produced with high efficacy in Heliothis virescens larvae. The capture-sandwich ELISA enabled a clear-cut qualitative evaluation of the CDV-specific immunoglobulin G (IgG) and IgM serostatuses of 196 and 35 dog sera, respectively. Inter-rater agreement analysis (kappa = 0.988) indicated that the ELISA can be used unrestrictedly as a substitute for the V-NA for the qualitative determination of CDV-specific IgG serostatus. In an attempt to semiquantify N-specific antibodies, a one-step-dilution (alpha method) IgG-specific ELISA was implemented. Alpha values of >/=50% showed very good inter-rater agreement (kappa = 0.968) with V-NA titers of >/=1/100 50% neutralizing dose (ND50) as measured against the central European CDV wild-type isolate 2544/Han95 in canine sera originating from northern Germany. An ND50 titer of 1/100 is considered a threshold, and titers of >/=1/100 indicate a resilient, protective immunity. CDV N-specific antibodies of the IgM class were detected by the newly developed ELISA in 9 of 15 sera obtained from dogs with symptoms of acute distemper. In leucocytes of 5 of the 15 dogs (all of which were also IgM positive) CDV RNA was detected by reverse transcription (RT)-PCR. The recombinant capture-sandwich ELISA detecting N-specific antibodies of the IgG class provided superior sensitivity and specificity and thus represents a rapid and cost-effective alternative to classical CDV V-NA. By detection of specific IgM antibodies, the ELISA will be complementary to RT-PCR and V-NA in the diagnosis of acute distemper infections. (+info)
Involvement of tachykinin receptors in sensitisation to cow's milk proteins in guinea pigs.
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BACKGROUND: There is growing evidence for a pivotal role for tachykinins in gut neuroimmune interactions. AIMS: To determine whether NK1, NK2, and NK3 tachykinin receptors are involved in milk protein induced allergic sensitisation. METHODS: Eight groups of 12 Dunkin-Hartley guinea pigs (250-300 g) were used. Four groups were sensitised to milk proteins for three weeks. During this period, these animals were injected intraperitoneally each day with NK1 (SR 140333; 0.3 mg/kg), NK2 (SR 48968; 5 mg/kg), or NK3 (SR 142801; 5 mg/kg) receptor antagonist or vehicle. The fifth group had water available instead of milk and was used as a non-sensitised control. The three other groups received the NK receptor antagonists for three weeks but were not sensitised to milk proteins. RESULTS: Sensitised animals treated with NK1 and NK3 receptor antagonists had both lower IgE and IgG serum titres, evaluated by passive cutaneous anaphylaxis, and lower specific IgG serum titres, determined by enzyme linked immunosorbent assay (ELISA), than vehicle treated animals. Sensitisation induced an increase in intestinal mast cell number which was abolished by treatment with the NK1 receptor antagonist. Antigenic challenge-induced jejunal hypersecretion was also blocked by treatment with the NK1 receptor antagonist. CONCLUSION: In guinea pigs, NK1 and NK3 but not NK2 receptors are involved in sensitisation to cow's milk. However, NK1 but not NK3 receptor antagonists abolish both the hypermastocytosis induced by food allergy and the hypersecretion induced by antigenic challenge, suggesting different roles for NK1 and NK3 receptors in the mechanisms of sensitisation to beta-lactoglobulin. (+info)
Prior cytomegalovirus infection and the risk of restenosis after percutaneous transluminal coronary balloon angioplasty.
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BACKGROUND: Restenosis is a common problem after all revascularization procedures in atherosclerotic coronary arteries. Reactivated human cytomegalovirus (CMV) has been detected in tissues of restenotic vascular lesions and was hypothesized to be a contributing pathogenic factor. Recent data suggest an association of restenosis after optimal coronary atherectomy with CMV serostatus, and a possible role of antiviral therapy was discussed. We therefore tested the hypothesis that prior CMV infection might be a risk factor for restenosis after conventional coronary balloon angioplasty (PTCA). METHODS AND RESULTS: We analyzed 92 consecutive patients who had been admitted for control angiography after previous PTCA within a mean interval of 6 months. Anti-CMV antibodies were measured as an indicator of prior CMV infection and latency. The coronary angiograms before PTCA, directly after, and 6 months later were analyzed quantitatively. Sixty-five percent of the patients were CMV-positive. Before PTCA, the degree (mean+/-SD) of stenosis was 69+/-10% in CMV-positive and 68+/-8.3% in CMV-negative subjects. PTCA resulted in a residual stenosis of 39% in both groups. After 6 months, the late losses of luminal diameter in the CMV-positive and -negative groups were 11+/-13% and 12+/-15%, respectively (P=0.658). In an ANCOVA with 25 potential risk factors for restenosis, CMV serostatus was not significantly associated with restenosis development. CONCLUSIONS: Our data indicate that prior CMV infection, in contrast to optimal atherectomy, is not associated with chronic restenosis after conventional coronary balloon angioplasty. The results do not support a possible benefit from antiviral therapy. (+info)
Natural history of Sin Nombre virus in western Colorado.
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A mark-recapture longitudinal study of immunoglobulin G (IgG) antibody to Sin Nombre virus (SNV) in rodent populations in western Colorado (1994-results summarized to October 1997) indicates the presence of SNV or a closely related hantavirus at two sites. Most rodents (principally deer mice, Peromyscus maniculatus, and pinyon mice, P. truei) did not persist on the trapping webs much beyond 1 month after first capture. Some persisted more than 1 year, which suggests that even a few infected deer mice could serve as transseasonal reservoirs and mechanisms for over-winter virus maintenance. A positive association between wounds and SNV antibody in adult animals at both sites suggests that when infected rodents in certain populations fight with uninfected rodents, virus amplification occurs. At both sites, male rodents comprised a larger percentage of seropositive mice than recaptured mice, which suggests that male mice contribute more to the SNV epizootic cycle than female mice. In deer mice, IgG antibody prevalence fluctuations were positively associated with population fluctuations. The rates of seroconversion, which in deer mice at both sites occurred mostly during late summer and midwinter, were higher than the seroprevalence, which suggests that the longer deer mice live, the greater the probability they will become infected with SNV. (+info)