Sputum Cytokine Levels in Patients with Pulmonary Tuberculosis as Early Markers of Mycobacterial Clearance

Sputum and serum from patients with active pulmonary tuberculosis (TB), healthy purified protein derivative-positive adults, and patients with bacterial pneumonia were collected to simultaneously assess local immunity in the lungs and peripheral blood. To determine whether cytokine profiles in sputum from TB patients and control subjects were a reflection of its cellular composition, cytospin slides were prepared in parallel and assessed for the presence of relative proportions of epithelial cells, neutrophils, macrophages, and T cells. Gamma interferon (IFN-γ) in sputum from TB patients was markedly elevated over levels for both control groups. With anti-TB therapy, IFN-γ levels in sputum from TB patients decreased rapidly and by week 4 of treatment were comparable to those in sputum from controls. Further, IFN-γ levels in sputum closely followed mycobacterial clearance. Although detected at fourfold-lower levels, IFN-γ immunoreactivities in serum followed kinetics in sputum. TNF-α, interleukin 8 (IL-8) and IL-6 also were readily detected in sputum from TB patients at baseline and responded to anti-TB therapy. In contrast to IFN-γ, however, TNF-α and IL-8 levels also were elevated in sputum from pneumonia controls. These data indicate that sputum cytokines correlate with disease activity during active TB of the lung and may serve as potential early markers for sputum conversion and response to anti-TB therapy.     

Salivary immunoglobulins in a patient with IgA deficiency

The saliva of an individual with selective IgA deficiency was found to contain IgG and IgM, with some of the IgM linked to secretory component. Some specimens showed evidence of low molecular weight immunoglobulin fragments, presumed to be the result of proteolysis.     

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.     

Chronic hepatitis C infection: influence of the viral load, genotypes, and GBV-C/HGV coinfection on the severity of the disease in a Brazilian population

The distributions of the different genotypes of the hepatitis C virus (HCV) and GBV-C virus (GBV-C/HGV) vary geographically and information worldwide is still incomplete. In particular, there are few data on the distribution of genotypes (and their relationship to the severity of liver disease) in South America. Findings are described in 114 consecutive patients from Northeast Brazil (median age 52 years, range 18-72 years) who had abnormal levels of serum aminotransferases and seropositivity for HCV RNA. The patients were recruited from an outpatient clinic between November 1997 and April 1998. Quantitative HCV RNA and GBV-C/HGV RNA estimations were carried out by double-nested polymerase chain reaction (PCR) using primers from the 5'-untranslated regions (UTRs) of the genomes. HCV genotypes were determined by restriction fragment length polymorphism (RFLP) analysis with 5'-UTR primers and by PCR with type-specific 5'-UTR primers. GBV-C/HGV-RNA genotypes were determined by RFLP with specific 5'-UTR primers and phylogenetic trees were constructed using the Neighbour-Joining and Drawtree programs. Histological features were graded and staged according to international criteria. Of the 114 patients, 35 (30.7%) patients had cirrhosis and 22 (27.8%) had mild, 51 (64.6%) had moderate, and 6 (7.6%) had severe chronic hepatitis. Median HCV viral load was 10(6) genome equivalents per millilitre (range 10(4)-10(9)/ml). Frequencies of genotypes were 5.3% type 1a, 44.7% type 1b, 3.5% type 2, 41.2% type 3, and 5.3% mixed types. GBV-C/HGV-RNA was detected in the sera of 12 (10.5%) patients and was distributed among three phylogenetic groups. There were no significant differences between patients with the predominant HCV genotypes (1b and 3) with respect to gender, age group, viral load, severity of liver disease, or coinfection with GBV-C/HGV.     

Immunohistochemical study of the expression of MUC5AC and MUC6 in breast carcinomas and adjacent breast tissues

AIM: To study the protein expression patterns of MUC5AC and MUC6 in normal and diseased breast tissues and to compare their expression with that of a mucin (MUC1) normally expressed in mammary tissues. METHODS: Formalin fixed, paraffin wax embedded tissue from 69 cases of invasive breast carcinoma and surrounding breast tissue was studied immunohistochemically with monoclonal antibodies against MUC1 (SM3), MUCSAC (CLH2), and MUC6 (CLH6), using the avidin-biotin-peroxidase method. RESULTS: MUC5AC was detected in five of 68 cases of invasive carcinoma including one of three cases of pure colloid carcinoma. MUCSAC expression in the adjacent normal breast epithelium was present in one of 29 cases and in one of two cases of ductal carcinoma in situ. None of 15 cases of ductal hyperplasia without atypia was positive for MUCSAC. MUC6 was present in 15 of 65 cases of invasive carcinoma, in four of 29 cases of normal adjacent epithelium, two of 15 cases of ductal hyperplasia without atypia, and one of two cases of ductal carcinoma in situ. MUC1 immunoreactivity detected by the SM3 antibody was present in 50 of the 67 cases of invasive carcinoma, but expression was also detected in benign epithelium. All invasive carcinomas expressing MUCSAC were positive for MUC1 and four were positive for MUC6. No significant association was found between the expression of these mucins and tumour size, histological grade, node status, oestrogen receptor status, p53 positivity, or c-ErbB-2 overexpression. CONCLUSIONS: This study documents the expression of two different mucins (MUCSAC and MUC6) not described as being expressed by normal breast tissues in a minority of breast carcinomas, as well as in normal and hyperplastic epithelium. Although the role of mucins in malignant transformation and the progression of breast cancer is not well understood, in some cases, there is probably an upregulation of several genes that encode distinct mucin proteins.     

Characterization of Paracoccidioides brasiliensis isolates by random amplified polymorphic DNA analysis.

We initially used 25 different random primers in order to test their ability to generate random amplified polymorphic DNA fragments from the dimorphic human pathogenic fungus Paracoccidioides brasiliensis. From the tested primers we chose five to distinguish between seven isolates of this microorganism. The DNA amplification patterns allowed clear differentiation of the seven isolates into two distinct groups with only 35% genomic identity. One of these groups contained two subgroups with 81% genetic similarity. The random amplified polymorphic DNA analysis method proved to be a good tool for analyzing and comparing different genomes of P. brasiliensis isolates.     

Histones interact with anionic phospholipids with high avidity; its relevance for the binding of histone-antihistone immune complexes.

Antibodies recognizing anionic phospholipids have been described in systemic lupus erythematosus (SLE) and other autoimmune diseases. Recent studies have shown that some of these antibodies may recognize a cardiolipin-binding protein (apolipoprotein H) rather than phospholipids. A similar possibility is conceivable for other cardiolipin-binding proteins that are targets of autoantibodies. In this study we have addressed whether this might be the case for histones, a set of highly cationic and widely distributed proteins that react in a well known autoantibody system. Our results indicate that: (i) histones bind to anionic phospholipids (cardiolipin and phosphatidylserine) with high avidity, but not to zwitterionic phospholipids (phosphatidylcholine); (ii) monoclonal and polyclonal antihistone antibodies recognize histones bound to cardiolipin; (iii) the addition of histones to serum samples containing antihistone antibodies often enhances their anticardiolipin reactivity. In addition, we have found that antihistone-producing hybridomas derived from MRL-lpr mice may show anticardiolipin activity due to the presence of histones in the cell culture supernatants with the resultant formation of immune complexes. Taken together, the results suggest a potential role for histones in the anti-cardiolipin activity detected in sera containing antihistone antibodies. These histone-phospholipid interactions should be taken into account when evaluating the pathogenic effects of antihistone antibodies or other autoantibodies reacting with nuclear components (e.g. nucleosomes) containing histones.