Atherosclerosis in survivors of Kawasaki disease.

OBJECTIVES: To test the hypothesis that long-term survivors of low-risk Kawasaki disease (KD) have ongoing vascular inflammation and dysfunction and a higher risk of accelerated atherosclerosis than healthy control subjects. STUDY DESIGN: Twenty-eight patients with KD (7-20 years after acute illness) and 27 age-matched healthy control subjects were examined for medical and dietary history, serum markers of atherosclerotic risk and inflammation, carotid intimal-medial thickness (CIMT) with vascular ultrasound scanning and arterial stiffness with applanation tonometry. RESULTS: Patients and control subjects were similar in age, sex, body mass index, waist-to-hip ratio, blood pressure, cigarette smoking, family history, diet, high-density lipoprotein cholesterol level, lipoprotein (a) level, homocysteine level, glucose level, insulin level, CIMT, arterial stiffness, C-reactive protein level, and inflammatory cytokine level. Levels of total cholesterol and apolipoprotein B were significantly higher in patients with KD than in control subjects. CONCLUSIONS: There was no evidence of increased atherosclerosis. Small but significant differences in cholesterol and apolipoprotein B levels could suggest increased future risk for atherosclerosis and warrant further study.

Comprehensive analysis of antibody responses to streptococcal and tissue antigens in patients with acute rheumatic fever.

Acute rheumatic fever (ARF) is an autoimmune disease occurring in individuals following untreated group A streptococcal infection believed to be triggered by antibodies to bacterial components that cross-react with human tissues. We developed a multiplexed immunoassay for the simultaneous quantitation of antibodies to nine streptococcal-related antigens including streptolysin O (SLO), DNase B, collagen I and IV, fibronectin, myosin, group A carbohydrate, M6 protein and streptococcal C5a peptidase. Utilizing this method, we examined serum from 49 ARF, 58 pharyngitis patients and age- and sex-matched controls in samples collected at initial disease onset, and at 4 weeks, 6 months and 1 year after diagnosis. Antibody responses were significantly higher for SLO, DNase B, M6 protein, group A carbohydrate and the cross-reactive antigens collagen I and myosin in ARF compared with pharyngitis patients (P

Identification of pyruvate kinase as an antigen associated with Tourette syndrome.

Immune responses to beta-hemolytic streptococcal infections are hypothesized to trigger tic disorders and early-onset obsessive-compulsive disorder (OCD) in some pediatric populations. Here we identify the M1 isoform of the glycolytic enzyme, pyruvate kinase (PK) as an autoimmune target in Tourette syndrome and associated disorders. Antibodies to PK reacted strongly with surface antigens of infectious strains of streptococcus, and antibodies to streptococcal M proteins reacted with PK. Moreover, immunoreactivity to PK in patients with exacerbated symptoms who had recently acquired a streptococcal infection was 7-fold higher compared to patients with exacerbated symptoms and no evidence of a streptococcal infection. These data suggest that PK can function as an autoimmune target and that this immunoreactivity may be associated with Tourette syndrome, OCD, and associated disorders.

D8/17 expression on B lymphocytes in anorexia nervosa.

OBJECTIVE: The authors' goal was to determine whether D8/17, a rheumatic fever susceptibility trait marker, identifies a possible type of anorexia nervosa: pediatric autoimmune neuropsychiatric disorders associated with streptococcus (PANDAS) anorexia nervosa. METHOD: Using immunofluorescence, the authors measured the percentage of D8/17-positive B lymphocytes in the peripheral blood of 16 subjects 7-21 years old who had not had rheumatic fever but who had possible PANDAS anorexia nervosa. The comparison subjects were 17 psychiatric patients with no eating disorder and no PANDAS characteristics. Subjects were considered D8/17 positive if they had 12% or more D8/17+ cells. RESULTS: There were more D8/17-positive individuals among those with PANDAS anorexia nervosa (81%) than among the comparison subjects (12%). The subjects with PANDAS anorexia nervosa had a higher percentage of D8/17+ cells (mean=27.1%, SD=17%) than the comparison subjects (mean=5.3%, SD=7.4%). CONCLUSIONS: A larger study is needed to determine whether D8/17 serves as a marker for susceptibility to a type of anorexia nervosa.

Studies of recombinant streptococcal pyrogenic exotoxin B/cysteine protease (rSPE B/SCP) in the skin of guinea pigs & the release of histamine from cultured mast cells & basophilic leukocytes.

BACKGROUND & OBJECTIVES: Streptococcal pyrogenic exotoxin B/streptococcal cysteine protease (SPE B/SCP) is considered to be one of the virulence factors of Streptococcus pyogenes (S. pyogenes) which causes serious diseases such as severe invasive infections and streptococcal toxic shock syndrome (STSS). There are no reports on the histamine releasing activity of SPE B/SCP from mast cells, although several biological activities have been studied. It is not clear whether SPE B/SCP have the superantigenic activity. We studied whether SPE B/SCP plays as a pathogenic factor in streptococcal infections and STSS through a histamine releasing activity. METHODS: Human mast cells and basophils were generated from CD34 positive cells isolated from cord blood and cultured in the presence of rIL-6, stem cell factor and/or rIL-3. The capacity of increasing capillary permeability of recombinant SPE B/SCP (rSPE B/SCP) was studied by using the skin of guinea pigs. Mitogenic activity to human T-cells of rSPE B/SCP was studied by incorporation of (3)Hthymidine. The levels of histamine in the plasma of patients with STSS and controls were measured by ELISA kit. RESULTS: rSPE B/SCP induced increased capillary permeability in the skin of guinea pigs, but both SPE A and SPE C did not exhibit such activity. Histamine was released from cultured human mast cells stimulated with rSPE B/SCP. The rSPE B/SCP did not exhibit mitogenic activity to human T-cells. Three of the 7 patients with STSS showed higher levels of plasma histamine than those of normal subjects. INTERPRETATION & CONCLUSION: The results suggested that increased capillary permeability and histamine release from mast cells induced by rSPE B/SCP might be involved in STSS and/or streptococcal infection of skin and mucous membrane.

Acute rheumatic fever and streptococci: the quintessential pathogenic trigger of autoimmunity.

Acute rheumatic fever (ARF) is a non-suppurative complication of pharyngeal infection with group A streptococcus. Signs and symptoms of ARF develop 2 to 3 weeks following pharyngitis and include arthritis, carditis, chorea, subcutaneous nodules, and erythema marginatum. In developing areas of the world, ARF and rheumatic heart disease are estimated to affect nearly 20 million people and remain leading causes of cardiovascular death during the first five decades of life. ARF still represents one of the quintessential examples of a pathogenic trigger culminating in autoimmune manifestations. In this review, we will focus on the pathogenesis and etiology of ARF and its complications, along with diagnostic and treatment approaches to both ameliorate and prevent long-term sequelae of this potentially debilitating disease.

Characterization of recombinant Streptococcus mitis-derived human platelet aggregation factor.

We previously purified Streptococcus mitis-derived human platelet aggregation factor (Sm-hPAF) from the culture supernatant of S. mitis strain Nm-65, isolated from the tooth surface of a patient with Kawasaki disease. Here we produced recombinant Sm-hPAF protein (rSm-hPAF) in Escherichia coli, to determine whether rSm-hPAF conserves its platelet aggregation activity. rSm-hPAF precursor (665 amino acids) shows up to 36-56% identity with the family of cholesterol-dependent cytolysins (CDCs), and rSm-hPAF displayed potent hemolytic activity toward mammalian erythrocytes, including human erythrocytes with platelet aggregation activity. The 162-amino acid amino-terminal domain of rSm-hPAF was found in no other CDCs except lectinolysin; this domain is homologous to a portion of pneumococcal fucolectin-related protein. Interestingly, suilysin (SLY) and pneumolysin (PLY) of CDCs also exhibit substantial human platelet aggregation activity, similar to rSm-hPAF, and the platelet aggregation by rSm-hPAF, SLY, and PLY was morphologically confirmed using light and electron microscopy.

Presence of D8/17 B-cell marker in patients with poststreptococcal reactive arthritis.

An elevated expression of the alloantigen D8/17 on B lymphocytes has been previously proposed as a susceptibility marker in rheumatic fever. The aim of the study was to investigate the presence of the D8/17 marker on B lymphocytes in poststreptococcal reactive arthritis (PSRA). The study sample included 19 patients (15 boys, 4 girls; mean age 11.7 +/- 5.4 years) who were diagnosed with PSRA (mean age at diagnosis, 9.4 +/- 5.6 years) and 18 healthy controls (10 boys and 8 girls) matched for ethnic background. B-cell D8/17 expression was tested by flow cytometry assay using monoclonal antibodies. Laboratory results showed a higher expression of D8/17 in the patient group (23.1 +/- 10.4%, range 11.6-51.9%) than in the control group (17.1 +/- 8.2%, range 7.1-35.7) in controls; this difference was statistically significant after log transformation of the data (P = 0.035). The high rate of expression of the D8/17 marker in patients with PSRA suggests that RF and PSRA share the same genetic susceptibility.

Group A streptococcus (GAS) carbohydrate as an immunogen for protection against GAS infection.

Previous studies have shown that human serum containing anti-group A streptococcus carbohydrate (GAS CHO) antibodies were opsonic for different M protein-carrying serotypes. To investigate the role that anti-GAS CHO antibodies play in passive and active protection, mice were immunized subcutaneously or intranasally with GAS CHO conjugated to tetanus toxoid, and mortality and oral colonization were monitored after challenge with live GAS. Compared with control mice, immunized mice were significantly protected against systemic or nasal challenge with GAS. Furthermore, studies of serum samples and throat cultures from Mexican children revealed an inverse relationship between high serum titers of anti-GAS CHO antibodies and the presence of GAS in the throat. Anti-GAS CHO antibodies were also tested for cross-reactivity with human tissues and cytoskeletal proteins. No cross-reactivity was observed in either assay. The present study demonstrates that GAS CHO is both immunogenic and protective against GAS infections.

Co-localization of human herpes virus 6 and tissue plasminogen activator in multiple sclerosis brain tissue.

BACKGROUND: Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system of unknown etiology. Several viruses have been suggested as playing a role in the pathogenesis of MS. The aim of this study was to investigate the interrelationship of human herpesvirus 6 (HHV-6) and plasminogen activation at the cellular level in MS plaques. MATERIAL/METHODS: Brain tissue specimens obtained from autopsies of 15 patients with MS and 10 controls were studied immunohistochemically for HHV-6 and cytomegalovirus (CMV) antigen and tissue plasminogen activator (tPA) protein. The presence of Ebstein-Barr virus (EBV) EBER RNA was studied using RNA in situ hybridization. RESULTS: HHV-6 antigen was identified in the cells of 67% (10/15) of MS brain sections and 30% (3/10) of the control sections. All samples were negative for CMV antigen and all samples with intact RNA were negative for EBV EBER RNA as demonstrated by in situ hybridization. tPA expression was found to be increased in MS plaques compared with the control samples. Interestingly, in 5 MS samples both HHV-6 antigen and tPA stained clearly, compared with none in the controls, but HHV-6 and tPA only occasionally co-localized in the same cells. CONCLUSIONS: At the cellular level, HHV-6 and plasminogen activation seem to co-localize in MS.

Antibodies to highly conserved peptide sequence of staphylococcal and streptococcal superantigens in Kawasaki disease.

Superantigen-mediated disease such as toxic shock syndrome is seen in patients who have a weak antibody response to the antigen toxic shock syndrome toxin 1 (TSST-1). We hypothesized that there may be deficiency in antibody production to staphylococcal and streptococcal toxins in Kawasaki disease (KD) children. A peptide was constructed from the homologous portion of the staphylococcal enterotoxins (SE) and streptococcal pyrogenic enterotoxins (SPE), and antibodies to the peptide were made. The anti-peptide antibody immunoblotted several of the SE toxins and SPE toxins. Presence of the peptide antibodies was investigated via ELISA in the sera of acute KD (n = 30), convalescent KD (n = 12), control adults (n = 10), and children (n = 19). The mean anti-peptide antibodies were indistinguishable between control children and KD before treatment with immunoglobulin (P = 0.7) but rose significantly after therapy (P < 0.01). The adults had significantly higher antibodies than the KD, both acute and late (P < 0.0001) and the control children (P < 0.0001). Thus, KD patients do not have a defective serological response against toxins such as SPE/SE/TSST-1. Normal children have significantly lower antitoxin antibody levels to the toxins compared to the adults.

Decreased levels of cystatin C, an inhibitor of the elastolytic enzyme cysteine protease, in acute and subacute phases of kawasaki disease.

Elevation of tissue-destructive proteases has been reported in acute Kawasaki disease. Cystatin C is a naturally occurring inhibitor of elastolytic cysteine protease in humans. Serum cystatin C deficiency in human beings has been linked to atherosclerosis and aortic aneurysms. We investigated the serum levels of cystatin C during acute Kawasaki disease. Serum samples from 17 acute Kawasaki disease patients were collected before and after immunoglobulin therapy and also at a median of 17 days after the therapy. Eight adults and 10 children without intercurrent infections served as control patients. Children with Kawasaki disease prior to therapy had significantly lower levels of cystatin C compared to adults (p = 0.002) and control children (p = 0.001). The low levels persisted 1-106 days after the therapy. Compared to control children and adults, children with Kawasaki disease had significantly lower serum levels of cystatin C in the acute stage before immunoglobulin therapy and in the subacute phase after the immunoglobulin therapy.

Staphylococcus aureus express unique superantigens depending on the tissue source.

A comprehensive analysis of Staphylococcus aureus superantigen (SAG) genes was undertaken in isolates from a major hospital and compared with isolates from patients with toxic shock syndrome (TSS). Polymerase chain reaction (PCR) analysis included recently discovered SAGs. Staphylococcal enterotoxin (SE) G and SEI were uniquely expressed in genital isolates. Genital isolates were similar to TSS isolates, although the latter frequently expressed TSS toxin 1. Both had a high frequency of SEG/SEI and a high number of SAG genes per bacterium. Detection of an SAG gene by PCR correlated with positive results in functional assays for SAG activity. Levels of serum antibodies to SEG and SEI, but not to other superantigens, were higher in healthy women than in men and served as an independent measure of the higher frequency of exposure to SEG/SEI among women. Together, the data suggest a role for SEG/SEI or closely linked genes in the adaptation of S. aureus to the genital mucosa environment.

Cysteine protease activity and histamine release from the human mast cell line HMC-1 stimulated by recombinant streptococcal pyrogenic exotoxin B/streptococcal cysteine protease.

We constructed the expression vector pSK-SCP containing the streptococcal exotoxin B gene (spe b) which expressed protease activity. We showed that the recombinant streptococcal pyogenic exotoxin B/streptococcal cysteine protease (rSPE B/SCP) was secreted into the culture supernatant of the transformant and retained its SCP activity, which was equivalent to or greater than that of the naturally occurring molecule. The secreted rSPE B/SCP induced histamine release and degranulation of the human mast cell line HMC-1. This study may contribute to the understanding of the pathogenic role of SPE B/SCP in streptococcal infection and streptococcal toxic shock syndrome.