Differential expression of CD22 (Lyb8) on murine B cells

Previous studies have established the distribution, biochemistryand functional attributes of human CD22, a B cell-restrictedglycoprotein. Recently, molecular cloning of the murine CD22equivalent revealed this molecule to be the same as the previouslydescribed Lyb8 alloantigen. Using the anti-Lyb8 mAb Cy34.1.2,the present report documents the expression patterns of CD22within the murine B cell compartment. The results demonstratethat in the bone marrow, murine CD22 is absent on the surfaceof pro-B cells, pre-B cells and newly emerging lgM⁺ B cells.CD22 is present at a low density on immature IgM^hi B cells andfully expressed on mature recirculating B cells. In the periphery,murine CD22 is expressed at mature levels on all B cell subsetsincluding follicular, marginal zone, B1 and switched B cells.Further studies showed CD22 to be retained on activated murineB cells for extended periods. Finally, in combination with CD23and heat stable antigen, CD22 can be used to delineate the immaturesplenic B cells, and distinguish them from follicular and marginalzone cells. Together, the results demonstrate murine CD22 tobe a useful pan marker for all mature B cell subsets.     

Immunohistochemical localization of apolipoproteins A-I and B in human carotid arteries

Decreased plasma levels of apolipoprotein A-I (apo A-I) and increased plasma levels of apolipoprotein B (apo B) have been shown to correlate with increased risk of atherosclerosis. While many studies have investigated the plasma levels of these apolipoproteins with regard to their value as predictors of cardiovascular disease, comparatively little is known about their precise tissue localization in atherosclerotic plaques. The purpose of this study was to determine the tissue localization of apo A-I and apo B in atherosclerotic segments of human carotid arteries through the use of immunohistochemical techniques. With tissue samples obtained from surgery and autopsy, apo A-I and apo B were found to be present in atherosclerotic plaques and absent in normal arterial tissue. In the plaques, both apo A-I and apo B were found extracellularly, primarily in the lipid core, but also in connective tissue. In addition, both apo A-I and apo B were found intracellularly in foam cells. This similar intracellular and extracellular distribution of apo A-I and apo B was unexpected, in view of their differing associations with atherosclerosis.     

Absence of peripheral clonal deletion and anergy in immune responses of T cell-reconstituted athymic mice

Superantigens induce clonal deletion of reactive T cells in the thymus and clonal deletion and anergy in the periphery of euthymic mice. In this report we have assessed the ability of Staphylococcal enterotoxin B (SEB) to induce peripheral tolerance in nude mice reconstituted with normal, syngeneic T cells. Immunization of reconstituted nude mice with SEB resulted in lethal toxic shock in a large fraction of the animals. Such lethality was never observed in the normal donor mouse strain. Analysis of lymphokine production in response to SEB showed that reconstituted nude mice produced higher levels of interleukin-2 and tumor necrosis factor-α, but lower levels of interleukin-4, than euthymic control mice. Furthermore, SEB was unable to promote either clonal elimination or induction of anergy in the SEB-responsive peripheral T cells, despite the fact that reconstituted nude mice did produce high levels of corticosterone upon treatment with SEB. These results imply a lack of control over immune responses to superantigen in T cell-reconstituted athymic mice.     

Anti-mitochondrial flavoprotein autoantibodies of patients with myocarditis and dilated cardiomyopathy (anti-M7): interaction with flavin-carrying proteins, effect of vitamin B2 and epitope mapping

Vitamin B2 and flavin cofactors are transported tightly bound to immunoglobulin in human serum. We reasoned that anti-mitochondrial flavoprotein autoantibodies (alpha Fp-AB) present in the serum of patients with myocarditis and cardiomyopathy of unknown aetiology may form immunoglobulin aggregates with these serum proteins. However, immunodiffusion and Western blot assays demonstrated that the flavin-carrying proteins were not recognized by alpha Fp-AB. Apparently the flavin moiety in the native protein conformation was inaccessible to alpha Fp-AB. This conclusion was supported by the absence of an immunoreaction between the riboflavin-binding protein from egg white and alpha FP-AB. Intravenous application of vitamin B2 to rabbits immunized with 6-hydroxy-D-nicotine oxidase, a bacterial protein carrying covalently attached FAD, did not neutralize alpha Fp-AB which had been raised in the serum of the animals. FAD-carrying peptides generated from 6-hydroxy-D-nicotine oxidase by trypsin and chymotrypsin treatment were not recognized by the alpha Fp-AB, but those generated by endopeptidase Lys were. This demonstrates that the epitope recognized by alpha Fp-AB comprises, besides the flavin moiety, protein secondary structure elements.     

A role for the VLA-4 integrin in the activation of human memory B cells

It is generally recognized that activation through membrane effector molecules such as CD40 or the B cell receptor (BCR) is mandatory to allow B cells to proliferate and differentiate into antibody (Ab)-secreting cells in response to cytokines. We show here that purified tonsillar B cells can be stimulated directly by a cytokine combination to proliferate and secrete immunoglobulins when cultures are performed at high cell density. The contact-mediated activation of B cells in this experimental system is strongly inhibited both by anti-very late antigen (VLA)-4 monoclonal Ab and by a peptide containing the LDV sequence specifically recognized by the α4 integrin binding site. These reagents also significantly suppressed the B cell responses elicited by engagement of the BCR or CD40. Our data reveal that memory B cells but not virgin or germinal center B cells are sensitive to the direct stimulatory effect of cytokines in high-density cultures. Finally, we found that the dual expression of the α and β chains of VLA-4 is a distinctive feature of the memory B cell population. Collectively, our findings support the notion that VLA-4-dependent homotypic B cell interactions can mediate a co-stimulatory signal to human memory B cells and might participate in the B cell activation triggered through the BCR and CD40.     

Attachment, invasion, chemotaxis, and proteinase expression of B16-BL6 melanoma cells exhibiting a low metastatic phenotype after exposure to dietary restriction of tyrosine and phenylalanine

We previously reported that low levels of tyrosine (Tyr) and phenylalanine (Phe) alter the metastatic phenotype of B16-BL6 (BL6) murine melanoma and select for tumor cell populations with decreased lung colonizing ability. To more specifically characterize the effects of Tyr and Phe restriction on the malignant phenotype of BL6, we investigated in vitro attachment, invasion, proteinase expression, and chemotaxis of high and low metastatic BL6 variants. High metastatic variant cells were isolated from subcutaneous tumors of mice fed a nutritionally complete diet (ND cells) and low metastatic variant cells were isolated from mice fed a diet restricted in Tyr and Phe (LTP cells). Results indicate that attachment to reconstituted basement membrane (Matrigel) was significantly reduced in LTP cells as compared to ND cells. Attachment to collagen IV, laminin, and fibronectin were similar between the two variants. Invasion through Matrigel and growth factor-reduced Matrigel were significantly decreased in LTP cells as compared to ND cells. Zymography revealed the presence of M _r 92 000 and M _r 72 000 progelatinases, tissue plasminogen activator, and urokinase plasminogen activator in the conditioned medium of both variants; however, there were no differences in activity of these secreted proteinases between the two variants. Growth of the variants on growth factor-reduced Matrigel similarly induced expression of the M _r 92 000 progelatinase. The variants exhibited similar chemotactic responses toward laminin. However, the chemotactic response toward fibronectin by LTP cells was significantly increased. MFR5, a monoclonal antibody which selectively blocks function of the ₅ chain of the ₅ß₁ integrin, VLA-5, decreased the chemotactic response toward fibronectin of ND cells by 37%; the chemotactic response by LTP cells was reduced by 49%. This effect was specific for fibronectin-mediated chemotaxis since the chemotaxis toward laminin and invasion through Matrigel were not altered by the presence of MFR5. The surface expression of VLA-5 was significantly increased in LTP cells as compared to ND cells by flow cytometric analysis. These observations suggest that limitation of Tyr and Phe either directly modifies BL6 or selects for subpopulations with altered in vitro invasion, chemotaxis, and integrin expression.     

Chronic mucocutaneous candidiasis. II. Class and subclass of specific antibody responses in vivo and in vitro

Patients with chronic mucocutaneous candidiasis (CMC) succumb to persistent infections with the opportunistic yeast Candida. Impaired cell-mediated responses to Candida have been repeatedly reported while antibody responses were mostly found to be normal. The underlying defect remains poorly understood. It has recently been shown that CMC patients are also susceptible to infections with encapsulated bacteria, and may have associated IgG2 and IgG4 deficiency. Our previous studies demonstrated altered cytokine production in CMC patients. As cytokines can influence production and isotype of specific antibody, in 10 patients with CMC we measured the levels and isotype distributions of serum antibodies to Candida antigens (CAg), pneumococcal polysaccharide (PPS) and tetanus toxoid (TT) antigens. Peripheral blood lymphocytes were also stimulated in culture and the antibodies made in vitro were measured. Our data demonstrated that in vivo, CMC patients had very high levels of IgG and IgA CAg-specific antibodies. CAg-specific and PPS-specific IgG1 was markedly higher than in controls. Children but not adults with CMC had significantly lower levels of IgG2-specific antibody to CAg and PPS compared with age-matched controls. Patients had significantly higher levels of IgG3-specific antibody to all three antigens tested. These findings were in accordance with increased total IgG and IgG3 levels seen in CMC patients. In vitro, CMC patients, particularly children, did not respond as frequently to antigen stimulation as did their healthy controls. The level of specific antibody produced was also lower to all antigens tested, as was the amount of total immunoglobulins following antigenic and particularly mitogenic stimulation. Addition of interferon-alpha (IFN-α) or IFN-γ to cultures had variable, sometimes marked, effects. Our results demonstrate that CMC patients manifest subtle alterations in specific antibody responses to CAg, PPS and TT, which are most pronounced in children. This may relate to altered cytokine production also seen in these patients.     

Environmental risk factors for hepatitis A infection in the Zenica–Doboj Canton, Bosnia and Herzegovina

In the Zenica-Doboj Canton, 1106 hepatitis A virus (HAV) infections were reported during 2000 (an incidence rate of 252/100 000 population), with 996 (90.1%) cases occurring in nine community-wide outbreaks. Analysis of water supplies showed that 398 (19.1%) samples contained coliforms, including 202 (50.8%) that were contaminated with thermotolerant Escherichia coli. Sewage sanitation systems were absent or substandard in 53 910 (81.8%) rural households. The group most affected during outbreaks comprised children aged 7-14 years (incidence rate of 598/100 000). The development of health promotion and prevention initiatives in schools, combined with rigorous hygiene measures, will be necessary to achieve control of the spread of HAV.     

Hepatitis E in the south west of France in individuals who have never visited an endemic area

A total of 431 consecutive patients from the Midi Pyrenees area with acute hepatitis with unknown etiology in 2001-2002 were tested for the presence of immunoglobulin G-class (IgG) anti-hepatitis E virus (HEV) antibodies. Forty-six (10.7%) had anti-HEV IgG, and the results were questionable for a further 17 (3.9%). Real time PCR based on TaqMan detection was used to identify HEV genome fragments in the serum of patients with positive or questionable anti-HEV serology. HEV RNA was found in 25.4% of cases. All amplification products were sequenced and analyzed. Phylogenetic analysis revealed that all the strains were genotype 3. In conclusion, virological and epidemiological data indicate that genotype 3 viruses are circulating in the south west part of France (Midi-Pyrenees) in patients with acute hepatitis and who have not visited recently areas in which HEV is endemic.     

Molecular epidemiological and clinical aspects of hepatitis D virus in a unique triple hepatitis viruses (B,C, D) endemic community in Taiwan

The molecular epidemiological and clinical aspects of hepatitis D virus (HDV) in a unique HBV, HCV, and HDV triple virus endemic community in southern Taiwan were investigated. A total of 2,909 residents aged 45 or older were screened for hepatitis B surface antigen (HBsAg), anti-HCV antibody, and anti-HDV antibody (specifically for HBsAg-positive carriers). Factors that might be associated with HDV infection, viral nucleic acid detection, and genotyping of HBV, HCV, and HDV were investigated. The prevalence of HBsAg and anti-HCV were 12.6% (366/2,909) and 41.6% (1,227/2,909), respectively. For HBsAg carriers, 15.3% (56/366) were positive for anti-HDV assay. Living in a higher endemic district of HCV infection (odds ratio [OR] = 3.2; 95% confidence interval [CI] = 1.7-6.3), male gender (OR = 1.9; 95% CI = 1.1-3.6) and co-infection with HCV (OR = 1.8; 95% CI = 1.0-3.3) were significantly independent factors associated with HDV infection. The detection rate of HDV RNA among anti-HDV-positive patients was only 12.7% (7/55). The mean HBV titer of triple infection group was significantly lower than in the HBV/HDV co-infection group (2.23 vs 3.05 in log(10), copies/ml, P = 0.046). HCV RNA detection among the triple infection group showed 47.4% (9/19) viremia rate and viral loads of 579,121 IU/ml in median (16,803-1,551,190 IU/ml). The prevalent genotype of HBV was type B (23/25); HCV was 1b (7/9) and HDV was IIa/IIb (4/4). Only the presence of HCV RNA predicted the presence of elevated ALT significantly (OR = 25.0; 95% CI = 3.39-184.6). In conclusion, the geographical aggregation of HDV infection paralleled that of HCV infection in this community. HCV suppressed the replication of HBV among triple vital infection patients. HBV and HDV lapsed into a remission or nonreplicative phase in most cases, and HCV acted as a dominant factor in triple viral-infected individuals. Only the presence of HCV RNA was associated with elevated ALT values, but not HBV or HDV.