Protein micelles of human histocompatibility antigens (HLA-A and HLA-B)

Human histocompatibility antigens (HLA-A and -B) are membrane proteins which have large hydrophilic domains outside the cell membrane and a small hydrophobic portion in the lipid bilayer. In this paper we describe optimal conditions for preparing micelles of detergent-solubilized HLA-A2 and -B7 antigens. These homogeneous protein aggregates are water soluble and free of detergent and lipid. Hydrophobic interactions between the intramembraneous portions of the HLA antigens are the driving forces in the formation of these protein micelles. The papain-solubilized fragment of the HLA antigens is not included in the micelle. The average molecular weight of the HLA micelles is around 9 × 10⁵ daltons, which suggests sixteen HLA-A2 and/or HLA-B7 antigenic molecules per protein aggregate. Electron microscopic studies revealed that the most frequent size of the micelles is 12 mm and that HLA-micelles are similar but not identical to micelles from Sindbis Virus glycoproteins (E1 and E2) The HLA-A2 and -B7 micelles retained full antigenic activity as judged by precipitations with allo- and heteroantisera. Such micelles will no doubt be important tools in further studies of the role of histocompatibility antigens.     

Nystatin studies of the skin of larval Rana catesbeiana

Transport and electrical characteristics of the isolated skin of larval Rana catesbeiana were analyzed using ion substitution and nystatin. When the inner (IBS) and outer (OBS) bathing solutions contained Na Ringer solution the electrical potential (TEP), short-circuit current (SCC), and resistance (R2) were 23.5 +/- 7.0 mV, 2.8 +/- 0.7 microA . cm-2, and 8.00 +/- 0.74 k omega. cm2, respectively (n = 4). When K was substituted for Na in the OBS these values were not changed significantly. When nystatin (120 U.cm-3), a drug that increases the permeability of membranes to small cations, was added to the OBS (Na Ringer) there was a striking increase in the TEP to 52.8 +/- 3.1 mV, SCC to 14.8 +/- 2.0 microA . cm-2, and drop in R2 to 3.75 +/- 0.52 k omega . cm2. The response to nystatin was similar with Na or K Ringer solution in the OBS (Na Ringer in the IBS). With Na Ringer in the OBS and IBS, the increase in SCC induced by low doses of nystatin equaled net Na flux measured isotopically. Plots of transepithelial conductance against SCC after nystatin were linear and provided estimates of shunt resistance (R*sh = 14.6 +/- 1.3 k omega . cm2) and electromotive driving force for ions (E*A = 76 +/- 3 mV). Similar curves were obtained with K Ringer in the OBS. In the presence of nystatin, characteristics of the basolateral membrane were evaluated. It displayed selective permeability to K relative to Na or Tris.     

Control of recombination within and between DNA plasmids of Saccharomyces cerevisiae

Summary [2 m⁺ and [2m°] yeast were transformed to stable leucine prototrophy with the hybrid yeast — E. coli plasmid, pJDB219. This plasmid contains the entire sequence of the endogenous 2 m yeast DNA plasmid in addition to the yeast nuclear LEU2 ⁺ gene and the Co1E1 derivative, pMB9. In the [2 m⁺] transformants, a new wholly yeast LEU2 ⁺ plasmid, pYX, was generated, probably by a recombination event between pJDB219 and 2 m DNA. The plamid, pYX, in the absence of 2 m DNA, was found to exist in equimolar amounts of two forms, A and B, which probably arise by intramolecular recombination across the inverted repeat sequences of the 2 m DNA portion of the plasmid. pJDB219 was found to require the presence of 2 m DNA to undergo this intramolecular recombination. The results suggest that 2, m DNA and pYX code for a gene product required in this recombination event which pJDB219 cannot produce.     

Genomic instability in scleroderma

Scleroderma is an enigmatic rheumatic disorder of uncertain etio-pathogenesis. Cancer has an approximately two-fold higher incidence in scleroderma patients than in the general population. There are preliminary data of acquired genetic damage in scleroderma but the significance of these observations are uncertain. To determine somatic mutation frequency at the glycophorin-A (GPA) locus in patients with limited and diffuse cutaneous scleroderma. The GPA assay measures the total somatic mutation frequency (Vf), composed of gene inactivating mutations (NO) and mutations arising from mitotic recombination (NN) in individuals heterozygous for the GPA MN blood group. Mutation frequency was determined using a validated GPA flow cytometric assay using fluorescent labeled monoclonal antibodies specific for the GPA blood groups M and N. This assay detects and enumerates progeny of red blood cell (rbc) precursor cells which have acquired genetic damage resulting in a loss of expression of one of the GPA alleles. It was found that patients with scleroderma (n = 23) had significantly elevated Vf as compared with young healthy controls (p < 0.001) and elderly controls (p = 0.03). Patients with diffuse scleroderma had higher mean Vf as compared with limited scleroderma (p = 0.055). In comparison with controls, patients with scleroderma exhibit a higher proportion of mitotic recombinant mutations than inactivating mutations (p < 0.002). There was no correlation between Vf and disease duration, age at onset or autoantibody status. We have documented evidence of acquired genetic damage at the GPA locus in scleroderma. Evidence of acquired genetic damage in this disorder may be importance in explaining both the etio-pathogenesis of scleroderma and the association of scleroderma with cancer.     

The sensitivity of antibody detection testing using pooled versus unpooled reagent red cells

Evaluation of paternity (alleged father, mother, and child) can range from a straightforward resolution to a complex problem that cannot be resolved without family studies. We present a case of disputed paternity in which tests for crossreactive groups (CREcS) and antigen subtypes (splits) within the human leukocyte antigen (HLA) system could not be used confidently to prove or disprove paternity. Further analysis, red cell enzyme tests, enabled a final verdict and confirmed the current reliability of HLA antisera defining splits.     

Psychotherapy and sociology: flirtation, marriage or divorce?

Psychotherapy and sociology are themselves products of mixed marriages; each experiences problems of identity and their children face problems of legitimacy. This paper considers the relevance of some sociological perspectives to the psychotherapeutic process, using the following analogies; (1) Flirtation: Why the attraction? 'going steady' or 'femme fatale'? (2) Marriage: Mutual enrichment, cross-fertilisation and new life? Examples of shared perspectives in the work of E. Becker, Erikson, Goffman etc. Influence of psychosocial contexts on psychotherapeutic processes. (3) Divorce: Too little shared life or too much destructive criticism? If the world within man and the world between men are interrelated, psychotherapy and sociology--although at times uneasy bedfellows--have much to give each other.     

Effects in mice of rat bromelain-treated RBC and lipopolysaccharide on autoantibody production against bromelain-treated isologous RBC

Autoantibody production against mouse bromelain-treated (brom) red blood cells (RBC) was significantly increased in mice injected with rat brom RBC. These autoantibodies were not adsorbed by rat brom RBC in serological assays and did not lyse rat brom RBC in plaque-forming cell (PFC) assays using mixtures of rat brom RBC and mouse brom RBC as targets. These data suggest that the increased response induced by rat brom RBC is not due to the presence of common, or similar, antigens on the two types of RBC. The spleens of mice injected with lipopolysaccharide (LPS), or with both LPS and rat brom RBC, had a markedly increased number of PFC lysing mouse brom RBC. About 20% of the PFC induced by LPS and rat brom RBC also lysed rat brom RBC. The autoimmune response was not increased in mice injected twice with rat brom RBC and the secondary response induced by two injections of LPS was lower than that induced by one injection of LPS. However, injection of LPS after an initial challenge with rat brom RBC induced an autoimmune response similar in size to that induced by LPS alone. The decreased secondary response against mouse brom RBC following a second injection of rat brom RBC was associated with decreased production of antibodies of various specificities as detected in a reverse PFC assay. These results do not support the hypothesis that the poor secondary responses against mouse brom RBC following a second injection of rat brom RBC are due to the exhaustive differentiation of autoimmune B cells as part of a fail-safe mechanism to prevent autoimmunity.