Biomedical subjects
Publications and source records attributed to J Scharfstein.
Amyloid P component deposition was detected by immunofluorescence with monospecific antibodies to human serum amyloid P in primary cell cultures of human smooth muscle cells isolated from blood vessels and gastric wall. Deposition was not detected in cultures of human skin fibroblasts.
Cells of Trypanosoma cruzi, Y strain, were submitted to water extraction by successive freezing and thawing. Fractionation of soluble material on a P-10 column gave an antigenic glycoprotein (fraction I) whose carbohydrate portion (40%) contained galactose, mannose, glucose, and xylose in a molar ratio of 35:13:1:1. It was electrophoretically homogeneous (Mr approximately 25 000) and contained short chains of mannopyranosyl (23%) and galactopyranosyl (10%) nonreducing end units and 2-O-substituted mannopyranosyl units (19%). Extraction of the remaining cell fragments with phenol-water gave an antigenic CHCl3-MeOH-H2O-soluble fraction II (LPPG), which yielded a galactomannan (fraction III) (galactose and mannose in a molar ratio of 1:2.1) on degradation with hot aqueous NaOH-NaBH4. It contained end units of galactofuranose (25%) and 2-O- (5%), 3-O- (32%), and 2,3-di-O- (19%) substituted mannopyranosyl units. Galactofuranose was directly linked (1 leads to 3) to mannopyranose and not via a phosphorodiester bridge. beta-D-Galf-(1 leads to 3)-Me-alpha-D-Manp, in contrast with Me-beta-D-Galf, was effective in inhibiting the precipitin reaction between LPPG and antiserum raised against LPPG. Fraction IV, insoluble in CHCl3-MeOH-H2O, contained galactose and mannose in a ratio of 1.4:1. After degradation with hot aqueous NaOH-NaBH4, it gave a product (fraction V) containing galactose and mannose in a 1:2 ratio. Methylation analysis showed it to differ from fraction III since it contained a high proportion of nonreducing end units (41%) and 2-O-substituted units (16%) of mannopyranose.
A glycoprotein of 25,000 daltons (G25) purified from T. cruzi extracts is recognized by serum antibodies of Chagas' disease patients. These human antibodies were isolated by affinity chromatography and were used to demonstrate that G25 antigenic determinants are i) represented at the parasite surface, and ii) are expressed in all developmental stages of the parasite's life cycle, as well as in several T. cruzi strains. This antigen-antibody system may be useful for the diagnosis of Chagas' disease because antibodies to radiolabeled G25 are found in the serum of 96.5% of 173 chagasic patients from different endemic areas, but are not found in the serum from other individuals. Taken collectively, the data suggest that antibodies to G25 define highly conserved determinants of the species T. cruzi. Moreover, its remarkable immunogenicity to infected humans offers an opportunity to investigate the role of specific immunologic responses in the pathogenicity of Chagas' disease.
Serum amyloid P protein (SAP), which shares several structural properties with C-reactive protein, has been recently identified as an acute-phase reactant in mice. In this study, the systemic inflammatory response of mice to infection with Trypanosoma cruzi was characterized with respect to induction of SAP as well as to stage-specific alterations on complement C3 and C4 levels. The SAP response depended on the dose and infectivity of the parasites. Kinetic data indicated a close temporal relationship between the onset of parasitemia and induction of SAP. The levels of SAP were maximally enhanced (1,050%) by the time parasitemia started to regress, and the response remained elevated as the infection entered the latency phase. The decline in parasitemia was paralleled by a significant reduction in C3 levels. A reciprocal relationship between the extent of parasitemia and SAP/C3 levels became apparent when these parameters were compared in individual inbred mice during the time of decreasing parasitemia.
C4-binding protein (bp), a glycoprotein with specific binding affinity for the activated form of C4 (C4b), has recently been isolated from human serum and partially characterized. This report demonstrates that C4-bp is incorporated into soluble immune complexes after complement activation in vitro. The reaction requires Ca++ ions and the presence of C4 in serum. Immunopathological studies of various forms of glomerulonephritis revealed intense C4-bp deposition in glomeruli from patients with immune-complex type of pathogenesis. C4-bp deposition was in close correlation with that of C4. These observations, together with the in vitro association of C4-bp to immune complexes, support the notion that the deposits in glomeruli represent the local accumulation of immune complexes.
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C4-binding protein (C4-bp), a new component of the complement system, was isolated from human plasma by precipitation with polyethyleneglycol, followed by chromatography on ion exchangers. C4-bp was identified on sodium dodecyl- sulfate polyacrylamide gel electrophoresis (SDS-PAGE) by two independent criteria: its ability to bind to C4b, and immunoprecipitation with a monospecificantiserum. Purified C4-bp is a 10.7 s glycoprotein. It consists of several disulfide bonded subunits of mol wt 70,000 daltons. Under nonreducing conditions, its mol wt has been estimated on SDS-PAGE as 540- 590,000 daltons. C4-bp moves as a slow B-globulin at pH 8.6 in the absence of free divalent cations, but when the buffers contain Ca(++)-lactate, C4-bp is a gamma globulin. Purified C4-bp binds to purified C4b. The reaction proceeds in the presence or absence of divalent cations and is not inhibited by diisopropylfluorophosphate. The C4b/C4-bp complexes have sedimentation coefficients between 15 and 17 s on sucrose gradient ultracentrifugation, and can be readily identified by crossed immunoelectrophoresis (CIE). The complexes move faster toward the anode than either protein. C4-bp is multivalent. Saturation is reached at molecular ratios of C4b/C4- bp of between 4 and 5. The interaction between C4b and C4-bp may complicate the electrophoretic patterns of these proteins in normal human serum, if the complement system is activated before or during the run. However, in EDTA-plasma, native C4 and C4-bp do not form stable complexes and can be identified in separate peaks after CIE.
In this paper, we sought to determine if chronic chagasic patients with cardiopathy could be distinguished from those displaying non-chagasic cardiopathy on the basis of T cell proliferative responses to cruzipain (GP57/51), a major antigen of T. cruzi. Assays were performed with peripheral blood mononuclear cells from 24 individuals classified as follows: normal donors (n = 8), patients with non-chagasic cardiopathy (n = 8), patients with chronic chagasic cardiopathy without morbid associations (n = 8). The analysis of variance indicated that the proliferative responses stimulated by cruzipain were significantly higher in the group of chagasic patients (p = 0.0001). Turkey's multiple comparison test showed that the proliferative index medium from normal and non-chagasic cardiopathy was not significantly different from each other. We conclude that the T cell responses against T. cruzipain, as measured by proliferative indices of cells found in peripheral blood, are exclusively associated with Chagas, disease. In view of the abundance of cruzipain antigen in amastigotes it is possible that these T cell specificities contribute to the heart tissue damage observed in chronic Chagas, disease patients.