Genetic control of endotoxic responses in mice.

A number of altered immunologic responses to lipopolysaccharide (LPS) in C3H/HeJ mice result from the expression in B lymphocytes of a defective genetic locus, termed Lps. Lps has been mapped to chromosome 4 between two loci, Mup-1 and Ps. As it is difficult to type individual mice for LPS responsiveness in more than one type of assay, we have utilized Mup-1 as a genetic marker to correlate LPS responses in mice to the expression of the Lps locus. Three nonlymphoid responses to LPS have been examined in 12 recombinant inbred strains of mice and in a backcross linkage analysis, and are all regulated by the expression of the Lps locus. These responses are hypothermal changes in body temperature, and the elevation in serum levels of a colony stimulating factor and the precursor of the secondary amyloid protein AA. Therefore, the initiation of LPS responses in different cell types in mice involve the expression of a common locus. These linkage studies provide a means for analyzing the genetic control of many of the diverse reactions of the endotoxic response to LPS.

Murine model for human secondary amyloidosis: genetic variability of the acute-phase serum protein SAA response to endotoxins and casein.

The serum precursor SAA of the secondary amyloid protein AA has been detected by solid-phase radioimmunoassay as a normal serum alpha-globulin of mol wt 160,000, which dissociates to a more stable 12,500 dalton moiety on treatment with formic acid. In 12 strains of mice, including T-cell-deficient nude mice, treated with the amyloid-inducing agents lipopolysaccharide (LPS) or casein, SAA behaved as an acute-phase reactant. SAA concentration rose to about 750 mug/ml by 24 h and returned to less than 1 mug/ml by 48 h. Since the amyloid-resistant colchicine-treated mice and AJ mice had a normal SAA response to LPS, it appears that their resistance to amyloid induction is due to the nature of their SAA processing rather than decreased SAA production. C3H/HeJ mice, which have defective B-lymphocyte responses to LPS, required extremely high dosages of LPS to cause SAA elevation, although their SAA response to casein was normal. This suggests that SAA is an acute-phase protein produced as a result of B-lymphocyte stimulation. Preliminary evidence suggests that at the height of an acute SAA response, liver homogenates are particularly rich in protein AA cross-reacting material.

Heterogeneity of human serum amyloid A proteins.

Serum amyloid A proteins (SAA), presumed precursors of the tissue amyloid A proteins (AA) characteristic of secondary amyloidosis, have been isolated from the plasma high-density lipoproteins (HDL) of normals after etiocholanolone-induced inflammation and from patients with Wegener's granulomatosis, systemic lupus erythematosis, juvenile rheumatoid arthritis, Waldenström's macroglobulinemia, and Goodpasture's syndrome. At least six polymorphic forms of SAA wer identified among the low molecular weight proteins of HDL, and these comprosed up to 27% of the total HDL protein. Gel and ion-exchange chromatography permitted isolation of the SAA polymorphs in homogeneous form. Their amino acid compositions were very similar, they were indistinguishable in cationic and sodium dodecyl sulfate-polyacrylamide gel electrophoresis systems, and each had the terminal sequency COOH-Tyr-Lys-Phe-. Charge heterogeneity in anionic-urea polyacrylamide gel electropherograms was unaffected by neuaminidase treatment, and none of the SAA protein bands stained with the periodate-Schiff reagent. The two major SAA polymorphs, designated SAA4 and SAA5 according to their order of elution from DEAE-cellulose, had different NH2-terminal sequences. Manual Edman degradation demonstrated NH2-arg-ser-phe-phe- for SAA4 and NH2-ser-phe-phe- for SAA5. This NH2-terminal heterogeneity corresponds to that most frequently reported for AA and suggests that microheterogeneity in SAA may underlie that already documented in AA. Sufficient quantitites of the other SAA polymorphs were not available for similar analyses, but the amino acid compositions do not indicate that NH2-terminal heterogeneity accounts for all of the observed polymorphism. Artifactual polymorphism also appears unlikely, and the heterogeneiy of SAA may reflect origin from more than one cell type with or without posttranslational modificaton. We calculate from quantitative COOH-terminal analyses that SAA is of 11,000-11,900 mol wt. Primary structure studies have shown AA t be a single chain protein of 76 residues, and SAA, therefore, appears to contain a peptide of 33 amino acids that is missing from AA.

Detection of a mediator derived from endotoxin-stimulated macrohpages that induces the acute phase serum amyloid A response in mice.

The mechanism by which LPS stimulates an acute phase serum amyloid A (SAA) response in C3H mice has been studied. A factor (SAA inducer) appears in the blood of C3H/HeN (lipopolysaccharide [LPS]-sensitive) mice approximately 1 h after administration of LPS, which, when passively administered, can induce C3H/HeJ mice to produce SAA although they are resistant to the LPS itself. SAA inducer has been detected in the culture medium of LPS treated C3H/HeN macrophages but not spleen cells. Thus, two stages in the induction of the acute phase SAA response are now recognized: a latent period of 2-3 h during which the SAA concentration remains at baseline values and in which SAA inducer appears, and the period of synthesis of SAA which lasts for approoximately 24 h past induction. It is proposed that a macrophage response to LPS is responsible for production of the serum mediator which induces SAA synthesis.

Changes in human serum amyloid A and C-reactive protein after etiocholanolone-induced inflammation.

Secondary amyloidosis is a complication of diseases characterized by recurrent acute inflammation. In this study, a standardized stimulus which induced fever and inflammation was given to six normal subjects (19-24 yr old) to follow the fluctuation in concentration of serum amyloid A (SAA), the precursor of the secondary amyloid fibril protein. After a single intramuscular injection of etiocholanolone (0.3 mg/kg), blood samples were drawn twice a day for 12 days for determination of SAA by solid phase radioimmunoassay. From a base line of <100 mug/ml, the SAA concentration began rising within 12 h to a maximum value at about 48 h of 1,350-1,800 mug/ml in three males and 380-900 mug/ml in three females and returned to base line by 4-5 days. The SAA response showed a similar time response to C-reactive protein (CRP), a well-documented acute phase protein which was assayed semiquantitatively by capillary tube precipitin reaction. CRP, but not SAA, showed a quantitative correlation with the amount of fever induced by etiocholanolone. One subject exhibited a second rise in SAA and CRP concentrations after acute over-indulgence with alcohol, suggesting that acute liver damage may have caused an acute phase reaction. Thus, a controlled episode of fever and inflammation produced a prompt and prolonged elevation of SAA and CRP concentrations. Unlike SAA, CRP has not been implicated in the pathogenesis of amyloidosis, although its relationship to the P component of amyloid has recently been established.

Efficacy of nine-valent pneumococcal conjugate vaccine against pneumonia and invasive pneumococcal disease in The Gambia: randomised, double-blind, placebo-controlled trial.

BACKGROUND: Pneumonia is estimated to cause 2 million deaths every year in children. Streptococcus pneumoniae is the most important cause of severe pneumonia. We aimed to assess the efficacy of a nine-valent pneumococcal conjugate vaccine in children. METHODS: We undertook a randomised, placebo-controlled, double-blind trial in eastern Gambia. Children age 6-51 weeks were randomly allocated three doses of either pneumococcal conjugate vaccine (n=8718) or placebo (8719), with intervals of at least 25 days between doses. Our primary outcome was first episode of radiological pneumonia. Secondary endpoints were clinical or severe clinical pneumonia, invasive pneumococcal disease, and all-cause admissions. Analyses were per protocol and intention to treat. FINDINGS: 529 children assigned vaccine and 568 allocated placebo were not included in the per-protocol analysis. Results of per-protocol and intention-to-treat analyses were similar. By per-protocol analysis, 333 of 8189 children given vaccine had an episode of radiological pneumonia compared with 513 of 8151 who received placebo. Pneumococcal vaccine efficacy was 37% (95% CI 27-45) against first episode of radiological pneumonia. First episodes of clinical pneumonia were reduced overall by 7% (95% CI 1-12). Efficacy of the conjugate vaccine was 77% (51-90) against invasive pneumococcal disease caused by vaccine serotypes, 50% (21-69) against disease caused by all serotypes, and 15% (7-21) against all-cause admissions. We also found an efficacy of 16% (3-28) against mortality. 110 serious adverse events arose in children given the pneumococcal vaccine compared with 131 in those who received placebo. INTERPRETATION: In this rural African setting, pneumococcal conjugate vaccine has high efficacy against radiological pneumonia and invasive pneumococcal disease, and can substantially reduce admissions and improve child survival. Pneumococcal conjugate vaccines should be made available to African infants.

NH2-terminal analysis of four of the polymorphic forms of human serum amyloid A proteins.

We recently demonstrated that the serum amyloid A proteins (SAA) occur in six related polymorphic forms of indistinguishable molecular weights and COOH-terminal sequence. We have now obtained very homogeneous preparations of four of these proteins and shown that their amino acid compositions are similar but not identical. Two of these, SAA1 and SAA4, have the same 20 NH2-terminal residues despite striking differences in electrophoretic mobility and solution properties. SAA5 and SAA2, respectively, lack one and three of the NH2-terminal residues common to SAA1 and SAA4. The data are consistent with the postulate that some of the SAA polymorphs are products of different genes.

Unanticipated amyloidosis in dogs infused with insulin.

Highly purified regular porcine insulin was given by portable insulin pumps through indwelling vena caval catheters to 17 (13 normal, and 4 pancreatectomized) dogs initially weighing 15 +/- 2 kg at rates ranging from 2 to 10 mU/min (total 17-250 mg) over time periods ranging from 37 to 252 days. During the course of the study, many of the animals lost weight and became anemic. Since these conditions persisted and weight loss progressed even after cessation of insulin infusion, as many of the dogs as possible (15 of 17) were autopsied for microscopic studies. Large amounts of amyloid were demonstrated in the liver, kidney, spleen, and/or pancreas in 55% (6/11) of normal, and in 75% (3/4) of pancreatectomized dogs. The amyloid deposits were Congo red positive, exhibited classical apple green fluorescence under polarized light, and possessed the characteristic ultrastructural features of amyloid. Massive deposits of amyloid were observed in animals receiving as little as 17 mg of insulin over a time span of 52 days. In those animals with hepatic amyloid, marked hepatomegaly was present (i.e., 1200 +/- 250, X +/- SD, versus 300 +/- 25 g for normal animals) and preterminal serum alkaline phosphatase levels were markedly elevated (434 +/- 285 versus 30 +/- 14 IU/L for animals without hepatic amyloid). The magnitude of the hepatic amyloid deposits precludes the possibility that they represent insulin aggregates or insulin-derived products per se. No evidence of amyloid was present in any of the tissue biopsy specimens obtained prior to insulin infusion. Moreover, the possibility that this represents an immune response to the injected porcine insulin has to be viewed in light of the fact that the amino acid sequences of dog and porcine insulins are identical. It is of particular interest that the affinity of the amyloid deposits for Congo red stain was totally abolished by prior permanganate treatment, suggesting that the amyloid was derived from serum amyloid A protein rather than from immunoglobulin light chains or insulin aggregates per se. Further evidence that the protein was of the AA-type came from the initial biochemical characterization. Gel filtration on Sephadex G100 in 6 M guanidine hydrochloride identified two small molecular weight peaks of about 13,000 and 25,000 daltons, both of which inhibited the radioimmunoassay for human AA protein.(ABSTRACT TRUNCATED AT 400 WORDS)

Inhibition of human neutrophil and Pseudomonas elastases by the amyloid P-component: a constituent of elastic fibers and amyloid deposits.

The amyloid P-component (AP), a ubiquitous component of amyloid fibrils, is also a plasma protein and a connective tissue constituent. Its proximity to elastin, in particular, suggested that AP might serve to protect elastic tissue from hydrolytic enzymes. The inhibition of pancreatic elastase by AP has been reported. In the present study, the effects of AP on human neutrophil elastase and Pseudomonas elastase were investigated, and AP was shown to interfere with the cleavage of soluble elastin. As indicated by Michaelis-Menten analysis, AP is acting as a noncompetitive inhibitor. C-reactive protein, which is structurally similar to AP, had no effect on either elastase. AP was also found to inhibit the degradation of secondary amyloid fibrils by neutrophil elastase when these structures were first partially purified and then reexposed to AP. AP's ability to inhibit elastase was compared with alpha-1 antitrypsin in the presence and absence of oxidizing agents. These substances, which are released by inflammatory cells, are known to abrogate alpha-1 antitrypsin's anti-protease capacity. This contributes to elevated levels of free proteases in the circulation and extravascular spaces during severe inflammation. AP is not susceptible to oxidation and remains a functional inhibitor under these conditions. The potential role of AP as an elastase inhibitor is discussed.

Relapse and mortality among HIV-infected and uninfected patients with tuberculosis successfully treated with twice weekly directly observed therapy in rural South Africa.

OBJECTIVE: To determine post-treatment relapse and mortality rates among HIV-infected and uninfected patients with tuberculosis treated with a twice-weekly drug regimen under direct observation (DOT). SETTING: Hlabisa, South Africa. PATIENTS: A group of 403 patients with tuberculosis (53% HIV infected) cured following treatment with isoniazid (H), rifampicin (R), pyrazinamide (Z) and ethambutol (E) given in hospital (median 17 days), followed by HRZE twice weekly to 2 months and HR twice weekly to 6 months in the community under DOT. METHODS: Relapses were identified through hospital readmission and 6-monthly home visits. Relapse (culture for Mycobacterium tuberculosis) and mortality given as rates per 100 person-years observation (PYO) stratified by HIV status and history of previous tuberculosis treatment. RESULTS: Mean (SD) post-treatment follow-up was 1.2 (0.4) years (total PYO = 499); 78 patients (19%) left the area, 58 (14%) died, 248 (62%) remained well and 19 (5%) relapsed. Relapse rates in HIV-infected and uninfected patients were 3.9 [95% confidence interval (CI) 1.5-6.3] and 3.6 (95% CI 1.1-6.1) per 100 PYO (P = 0.7). Probability of relapse at 18 months was estimated as 5% in each group. Mortality was four-fold higher among HIV-infected patients (17.8 and 4.4 deaths per 100 PYO for HIV-infected and uninfected patients, respectively; P<0.0001). Probability of survival at 24 months was estimated as 59% and 81%, respectively. We observed no increase in relapse or mortality among previously treated patients compared with new patients. A positive smear at 2 months did not predict relapse or mortality. CONCLUSION: Relapse rates are acceptably low following successful DOT with a twice weekly rifampicin-containing regimen, irrespective of HIV status and previous treatment history. Mortality is substantially increased among HIV-infected patients even following successful DOT and this requires further attention.

Twice-weekly, directly observed treatment for HIV-infected and uninfected tuberculosis patients: cohort study in rural South Africa.

OBJECTIVE: To determine the effectiveness of twice-weekly directly observed therapy (DOT) for tuberculosis (TB) in HIV-infected and uninfected patients, irrespective of their previous treatment history. Also to determine the predictive value of 2-3 month smears on treatment outcome. METHODS: Four hundred and sixteen new and 113 previously treated adults with culture positive pulmonary TB (58% HIV infected, 9% combined drug resistance) in Hlabisa, South Africa. Daily isoniazid (H), rifampicin (R), pyrazinamide (Z) and ethambutol (E) given in hospital (median 17 days), followed by HRZE twice a week to 2 months and HR twice a week to 6 months in the community. RESULTS: Outcomes at 6 months among the 416 new patients were: transferred out 2%; interrupted treatment 17%; completed treatment 3%; failure 2%; and cured 71%. Outcomes were similar among HIV-infected and uninfected patients except for death (6 versus 2%; P = 0.03). Cure was frequent among adherent HIV-infected (97%; 95% CI 94-99%) and uninfected (96%; 95% CI 92-99%) new patients. Outcomes were similar among previously treated and new patients, except for death (11 versus 4%; P = 0.01), and cure among adherent previously treated patients 97% (95% CI 92-99%) was high. Smear results at 2 months did not predict the final outcome. CONCLUSION: A twice-weekly rifampicin-containing drug regimen given under DOT cures most adherent patients irrespective of HIV status and previous treatment history. The 2 month smear may be safely omitted. Relapse rates need to be determined, and an improved system of keeping treatment interrupters on therapy is needed. Simplified TB treatment may aid implementation of the DOTS strategy in settings with high TB caseloads secondary to the HIV epidemic.

The impact of HIV on infectiousness of pulmonary tuberculosis: a community study in Zambia.

OBJECTIVE: To examine the impact of HIV on infectiousness of pulmonary tuberculosis (TB). DESIGN: A cross-sectional tuberculin survey carried out among household contacts of HIV-1-positive and negative patients with bacteriologically confirmed pulmonary TB. Contacts were also examined for active TB. SETTING: Index cases were recruited from patients attending the University Teaching Hospital in Lusaka, Zambia and household contacts were examined during visits to their homes within Lusaka. PATIENTS, PARTICIPANTS: A total of 207 contacts of 43 HIV-positive patients, and 141 contacts of 28 HIV-negative patients with pulmonary TB were examined. MAIN OUTCOME MEASURES: Proportion of contacts of HIV-positive and negative index cases with a positive tuberculin response (diameter of induration > or = 5 mm to a dose of 2 tuberculin units). RESULTS: Fifty-two per cent of contacts of HIV-positive pulmonary TB patients had a positive tuberculin response compared with 71% of contacts of HIV-negative patients (odds ratio, 0.43; 95% CI, 0.26-0.72; P < 0.001). This difference persisted after allowing for between-household variations in the tuberculin response. Tuberculin response in the contact was related to age of contact, intimacy with the index case and crowding in the household. However, the effect of HIV status of the index case was not confounded by these variables. Tuberculin response in the contact was also related to the number of bacilli seen in the sputum smear of the index case which partially explained the effect of HIV status of the index case. Active TB was diagnosed in 4% of contacts of HIV-positive and 3% of contacts of HIV-negative cases, respectively (P = 0.8). CONCLUSIONS: HIV-positive patients with pulmonary TB may be less infectious than their HIV-negative counterparts and this may partly be explained by lower bacillary load in the sputum.

PIP: Between April and December 1989, the chest clinic of the University Teaching Hospital in Lusaka, Zambia, confirmed pulmonary tuberculosis (TB) in 141 adults, 95 (67%) of whom were HIV-1 seropositive. Health workers made home visits to 71 of the index cases (43 HIV-1 positive and 28 HIV-1 negative) to learn whether the 348 household members would also develop TB, thus allowing researchers to determine the effect of HIV on infectiousness of TB. Contacts of HIV-1 positive patients developed TB at a lower rate than did those of HIV-1 negative patients (52% vs. 71%; odds ratio [OR] = 0.43; p .001). This difference continued even after controlling for between-household variations, indicating that confounding variables did not account for the difference. Age of contact, intimacy with the index case, and crowding in the household were associated with the tuberculin response in the contact, but they did not confound the effect of HIV status. Tuberculin response in the contact was associated with the number of bacilli in the sputum smear (crude OR = 3.13; p = .013, and adjusted OR =1.84; p = .28), suggesting that the number of bacilli somewhat explained the difference in infectiousness between HIV-1 positive and HIV-1 negative patients. 12 contacts (8 of HIV-positive cases and 4 of HIV-negative cases) developed active TB after the TB diagnosis in the index case. These findings clearly demonstrated that infection with Mycobacterium tuberculosis was less likely in household members of HIV-1 positive cases than in those of HIV-1 negative cases. The lower bacillary load in the sputum in HIV- 1 cases may have accounted somewhat for the lower infectiousness of pulmonary TB.

Twice weekly tuberculosis preventive therapy in HIV infection in Zambia.

BACKGROUND: A randomized double-blind placebo-controlled trial was conducted to estimate the efficacy of preventive therapy for tuberculosis (TB) in HIV-infected adults in Lusaka, Zambia. The main outcome measures were the incidence of TB, mortality and adverse drug reactions. METHODS: During a 2 year period, 1053 HIV-positive individuals without evidence of clinical TB were randomly assigned to receive 6 months of isoniazid twice a week (H), or 3 months of rifampicin twice a week (R) plus pyrazinamide (Z), or a placebo. Therapy was taken twice a week and was self administered. Subjects presenting with symptoms during the follow-up period were investigated for TB. RESULTS: The 1053 subjects in the study were followed up for a total of 1631 person-years (median = 1.8 years). Twenty-nine subjects were taken off treatment as a result of adverse drug reactions. A total of 96 cases of TB/probable TB (59 TB and 37 probable TB) were diagnosed during the study period and 185 deaths were reported. One hundred and fifteen subjects (11%) did not return to the study clinic at any time after enrolment. The incidence of TB was lower in those subjects on preventive therapy (H and RZ groups combined) compared with those on placebo (rate ratio = 0.60, 95% CI: 0.36-1.01, P = 0.057), as was the incidence of TB/probable TB (rate ratio = 0.60, 95% CI: 0.40-0.89, P = 0.013). The effect of preventive therapy was greater in those with a tuberculin skin test (TST) of 5 mm or greater, in those with a lymphocyte count of 2x10(9)/l or higher, and in those with haemoglobin of 10 g/dl or higher. There was no difference in mortality rates between the preventive therapy and placebo groups. The effect of preventive therapy declined after the first year of the study so that by 18 months the rates of TB in the treated groups were similar to that in the placebo group. CONCLUSION: This study has demonstrated that preventive therapy with either twice weekly isoniazid for 6 months or a combination of rifampicin and pyrazinamide for 3 months reduced the incidence of TB in HIV-infected persons in Zambia. No effect was observed on mortality. The effect was greatest in persons who had a positive TST or a lymphocyte count of 2x10(9)/l or greater, indicating that preventive therapy may be more effective in people with less advanced immunosuppression. The limited duration of the protective effect reported in this study raises the question of the need for lifelong preventive therapy or re-prophylaxis.

Isoniazid preventive therapy for tuberculosis in HIV-1-infected adults: results of a randomized controlled trial.

OBJECTIVES: To determine the efficacy of isoniazid 300 mg daily for 6 months in the prevention of tuberculosis in HIV-1-infected adults and to determine whether tuberculosis preventive therapy prolongs survival in HIV-1-infected adults. DESIGN AND SETTING: Randomized, double-blind, placebo-controlled trial in Nairobi, Kenya. SUBJECTS: Six hundred and eighty-four HIV-1-infected adults. MAIN OUTCOME MEASURES: Development of tuberculosis and death. RESULTS: Three hundred and forty-two subjects received isoniazid and 342 received placebo. The median CD4 lymphocyte counts at enrolment were 322 and 346 x 10(6)/l in the isoniazid and placebo groups, respectively. The overall median follow-up from enrolment was 1.83 years (range, 0-3.4 years). The incidence of tuberculosis in the isoniazid group was 4.29 per 100 person-years (PY) of observation [95% confidence interval (CI) 2.78-6.33] and 3.86 per 100 PY of observation (95% CI, 2.45-5.79) in the placebo group, giving an adjusted rate ratio for isoniazid versus placebo of 0.92 (95% CI, 0.49-1.71). The adjusted rate ratio for tuberculosis for isoniazid versus placebo for tuberculin skin test (TST)-positive subjects was 0.60 (95% CI, 0.23-1.60) and for the TST-negative subjects, 1.23 (95% CI, 0.55-2.76). The overall adjusted mortality rate ratio for isoniazid versus placebo was 1.18 (95% CI, 0.79-1.75). Stratifying by TST reactivity gave an adjusted mortality rate ratio in those who were TST-positive of 0.33 (95% CI, 0.09-1.23) and for TST-negative subjects, 1.39 (95% CI, 0.90-2.12). CONCLUSIONS: Overall there was no statistically significant protective effect of daily isoniazid for 6 months in the prevention of tuberculosis. In the TST-positive subjects, where reactivation is likely to be the more important pathogenetic mechanism, there was some protection and some reduction in mortality, although this was not statistically significant. The small number of individuals in this subgroup made the power to detect a statistically significant difference in this subgroup low. Other influences that may have diluted the efficacy of isoniazid include a high rate of transmission of new infection and rapid progression to disease or insufficient duration of isoniazid in subjects with relatively advanced immunosuppression. The rate of drug resistance observed in subjects who received isoniazid and subsequently developed tuberculosis was low.

High level expression of genes cloned in phage lambda gt11.

Plasmid cloning vectors have been constructed which allow genes originally cloned in lambda gt11 to be expressed at a high level in Escherichia coli. They are based on the pEMBL and pUC vectors, with the genes transcribed from the lac promoter. The EcoRI site in the vector has been altered to be in the same reading frame as the site used for cloning in lambda gt11. Cloned proteins are expressed fused to a 2-kDa leader sequence containing a run of six Aparagine residues which considerably improves the stability of the recombinant proteins, but does not interfere with immunological assays. Using these vectors, the Mycobacterium leprae 18-kDa protein was expressed at 20 mg per litre of culture and constituted 15% of total cell protein.

Amyloidosis in systemic lupus erythematosus.

Amyloidosis occurs in a significant proportion of patients with rheumatologic diseases. The fibrillar amyloid proteins in such patients are composed predominantly of amyloid A protein, which is characteristic of the amyloid deposits associated with chronic inflammatory diseases. Only four patients with amyloidosis associated with systemic lupus erythematosus (SLE) have been described previously; analyses of their fibrillar amyloid proteins were not reported. We present herein, a patient with SLE and amyloidosis. Histochemical staining of our patient's renal tissue with Congo red demonstrated that the amyloid deposits contained amyloid A protein, as defined by permanganate sensitivity. In addition, the patient's serum contained increased concentrations of serum amyloid A proteins. In review, each of the previously described patients with amyloidosis associated with SLE had renal amyloid deposits, with diagnosis in three during evaluation of proteinuria. Thus, although rare, amyloidosis should be considered in the differential diagnosis of proteinuria in patients with SLE.

Decreased IFN- gamma and increased IL-4 production by human CD8(+) T cells in response to Mycobacterium tuberculosis in tuberculosis patients.

To investigate the role of MHC class I restricted CD8(+) T cells in host defense to M. tuberculosis, peripheral blood mononuclear cells (PBMC) from healthy BCG-vaccinated donors and untreated pulmonary tuberculosis (TB) patients in The Gambia were stimulated for 6 days with M. bovis BCG or M. tuberculosis and the CD8(+) T cell response analyzed. Intracellular FACS analysis of cytokine production by CD8(+) T cells showed that IFN- gamma and TNF- alpha production were greatly reduced in TB patients compared to healthy controls. IL-4-producing CD8(+) T cells were detected in TB patients, a phenotype absent in controls. Collectively, these data suggest that an alteration in the type 1/type 2 cytokine balance occurs in CD8(+) T cells during clinical tuberculosis, and that this may provide a surrogate marker for disease.

Amyloid cardiomyopathy: characterization by a distinctive voltage/mass relation.

Fourteen patients with biopsy-proved systemic amyloidosis underwent noninvasive cardiac testing to assess the presence and severity of cardiac amyloidosis. There was a clear tendency for electrocardiographic voltage to be low (sum of S wave in lead V1 plus R wave in lead V5 or V6 [SV1 + RV5 or V6] = 14.6 +/- 4.8 mm; normal range 15 to 35) and echocardiographic muscle cross-sectional area to be increased (11.4 +/- 2.7 cm2/m2; normal range 6 to 10). When the electrocardiographic or the echocardiographic data were examined individually, and especially when they were compared and contrasted with similar measurements from patients with pericardial disease (n = 8) or aortic valve disease (n = 24), it was apparent that the electrocardiogram and the echocardiogram had limited specificity in the diagnosis of amyloidosis. However, when the analysis combined these two techniques, a distinctive pattern emerged. There was an inverse correlation between voltage and muscle cross-sectional area (r = -0.79) in patients with amyloidosis; moreover, marked derangement of the voltage/cross-sectional area relation was associated with clinical symptoms and mortality. In addition, patients with amyloidosis and cardiac symptoms had abnormal left ventricular chamber radius to wall thickness ratios, consistent with infiltration of the myocardium as the primary abnormality in cardiac amyloidosis.

Human monoclonal antibodies to phenolic glycolipid-1 from leprosy patients cross react with poly(ADP-ribose), polynucleotides and tissue bound antigens.

Antibodies which bind to poly(ADP-ribose) have been described in Systemic Lupus Erythematosus (SLE) and a variety of infectious diseases. Two IgM kappa human monoclonal antibodies (MAbs), TH3 and PR4, produced from the fusion of peripheral blood lymphocytes of leprosy patients with the GM4672 lymphoblastoid cell line, were found to bind to poly(ADP-ribose) in direct binding and inhibition ELISAs. Significant inhibition of binding of these MAbs to poly(ADP-ribose) occurred with phenolic glycolipid-1, the M. leprae specific glycolipid, ssDNA, dsDNA, poly(dT), as well as poly(ADP-ribose) itself. Up to 80% of binding of TH3, and 90% of binding of PR4, to poly(ADP-ribose) was inhibited by 10 mcg of ssDNA suggesting that there may be sharing of some conformational determinants. Although the serological binding profiles of TH3 and PR4 are similar, only PR4 was found to bind to basal keratinocytes of normal human interfollicular epidermis and astrocyte cytoplasm in normal brain tissue. These results support the concept that an antibody binding site may accommodate more than one epitope. Furthermore, small differences in antigen binding potential may distinguish relatively innocuous antibodies from those which may be more pathogenic.

Diagnosis of Streptococcus pneumoniae pneumonia by quantitative enzyme linked immunosorbent assay of C-polysaccharide antigen.

AIMS: To evaluate the use of a quantitative enzyme linked immunosorbent assay (ELISA) detecting C-polysaccharide (PnC) antigen in sputum for the diagnosis of Streptococcus pneumoniae infection. METHODS: Specimens of sputum from 60 patients with acute community and hospital acquired pneumonia and infective exacerbations of obstructive airways disease were examined by semiquantitative culture and antigen ELISA. RESULTS: Using a cutoff value of 1 microgram/ml PnC antigen for a positive result, the sensitivity of this assay was 90.3%, specificity 93.1%, predictive value of a positive result was 93.5%, and the predictive value of a negative result 89.6%. CONCLUSIONS: Quantitation of C-polysaccharide antigen in sputum by ELISA distinguishes between carriage of oral bacteria which express PnC-like antigen and infection with S pneumoniae and compares favourably with other diagnostic methods.