MAM: a "new" high-incidence antigen found on multiple cell lines.

BACKGROUND: Three women have been identified with an antibody to a "new" high-incidence antigen found on multiple cell lines. CASE REPORTS: The proposita, M.A.M., presented during her third pregnancy with an antibody reacting with all RBCs tested except her own. She delivered a thrombocytopenic infant with a 3+ DAT, but without symptoms of HDN. The second example, A.N., presented during her third pregnancy with an antibody reacting with all RBCs tested except her own and those of M.A.M. She delivered a slightly thrombocytopenic but severely anemic infant. The third example, F.K., a sister of A.N., has an antibody reacting with all RBCs tested except her own and those of M.A.M. and A.N. CONCLUSION: This "new" high-incidence antigen has been named MAM and assigned high-incidence antigen number 901016 by the International Society of Blood Transfusion. The corresponding antibody, anti-MAM, has been shown to cause HDN and has the potential to shorten RBC survival after the transfusion of incompatible RBC units, as determined by monocyte monolayer assay. Immunoblotting and flow cytometry show that this new antibody reacts with various WBC lines in addition to RBCs. This antibody also appears to react with platelets in some assays.

Independent roles for platelet crossmatching and HLA in the selection of platelets for alloimmunized patients.

BACKGROUND: Although HLA-matched platelets are frequently requested for alloimmunized patients, recent evidence has indicated that 1-hour posttransfusion platelet increments in these patients are specifically sensitive to crossmatch compatibility. STUDY DESIGN AND METHODS: To determine the extent of advantage gained by use of single-donor apheresis (SD) platelets selected on the basis of HLA match when crossmatch-compatible SD platelets were available, a total of 220 platelet transfusions given in the absence of individually determined significant nonimmune factors were analyzed in a well-characterized cohort of platelet-refractory patients. Platelets were selected by solid-phase crossmatch from a small donor pool of relatively poor HLA matches or, upon request, ordered as HLA-matched and later crossmatched. RESULTS: Alloimmunized patients responded better to SD platelets selected on the basis of HLA than to pooled platelet concentrates or SD platelets selected at random, although most of the benefit was limited to the 57-percent subset of good HLA matches. Crossmatch-compatible SD platelets provided similar posttransfusion platelet increments independent of the HLA match. None of 31 crossmatch-incompatible SD platelets transfused provided an adequate increment, including 13 that were ordered as HLA-matched platelets. CONCLUSION: No benefit could be demonstrated from requesting that SD platelets be HLA-matched when crossmatch-compatible SD platelets were available.

Characterization of PKC2, a gene encoding a second protein kinase C isotype of Saccharomyces cerevisiae.

BACKGROUND: Protein kinase C (PKC) has attracted considerable attention over the past decade, primarily because of its presumed role in cellular growth control and tumourigenesis. Mammalian cells express at least 10 different isozymes of PKC; it is this complexity that has made elucidating the precise functions of PKC: so difficult. The identification of PKC homologues in organisms such as Drosophila, Xenopus, Dictyostelium, Aplysia and Caenorhabditis indicates that the enzyme is evolutionarily conserved, and this has stimulated our search for counterparts in the yeast Saccharomyces cerevisiae, in which powerful genetic analyses can be used. To date, only one PKC homologue, PKC1, has been identified in yeast and no biochemical activity has been definitively ascribed to the encoded protein. This, and the inability to identify other PKC homologues in yeast by DNA hybridization, has led to doubts about the existence of PKC isozymes in yeast. We have taken the approach of screening yeast expression libraries with anti-PKC antibodies in an attempt to identify further homologues. RESULTS: We have identified a novel PKC isozyme, Pkc2p, encoded by the gene PKC2. We report here the sequence of PKC2 and a comparison showing its similarity to other PKCs. Phylogenetic analysis suggests that all known PKC genes, including PKC2, originated from a common ancestor. Disruption of the PKC2 protein-coding region, deleting the entire catalytic domain of the encoded enzyme, is not lethal to yeast growing on rich media. However, the pkc2 mutant, unlike wild-type strains, fails to grow on minimal media containing limited concentrations of amino acids. This implicates Pkc2p in the response of yeast cells to amino-acid starvation. CONCLUSION: We have shown that yeast cells do express more than one PKC isozyme, by identifying and characterizing a novel PKC gene PKC2, the product of which may be involved in the cellular response to amino-acid starvation.

A proline-rich protein, verprolin, involved in cytoskeletal organization and cellular growth in the yeast Saccharomyces cerevisiae.

A gene (VRP1) encoding a novel proline-rich protein (verprolin) has been isolated from the yeast Saccharomyces cerevisiae as a result of its hybridization to a chick vinculin cDNA probe. The deduced protein sequence contains 24% proline residues present as proline-rich motifs throughout the verprolin sequence. Several of these motifs resemble recently identified sequences shown to bind Src homology 3 (SH3) domains in vitro. Replacement of the wild-type VRP1 allele with a mutant allele results in strains that grow slower than wild-type strains and are temperature sensitive. The vrp1 mutants are impaired in both cell shape and size and display aberrant chitin and actin localization. We propose that verporlin is involved in the maintenance of the yeast actin cytoskeleton, through interactions with other proteins, possibly containing SH3 domains.

Clinical and blood bank factors in the management of platelet refractoriness and alloimmunization.

Numerous independent and interdependent factors are involved in the posttransfusion platelet response. Factors such as ABO match and platelet age are related to circumstances potentially under the control of the blood bank physician and therefore may permit circumvention by an active transfusion service. On the other hand, factors such as fever or sepsis may be unavoidable, being related more to the individual patient or clinical condition. To evaluate which factors could be circumvented, we prospectively followed the 1-hour corrected count increments (CCIs) for 962 single-donor apheresis platelet transfusions to 71 refractory hematologic oncology inpatients, with concomitant recording of implicated factors. Stepwise regression analysis allowed for determination of which concurrent and confounding clinical-, patient-, and blood bank-related factors significantly affected the CCIs. Although many implicated factors proved to be independently associated with an increased or decreased CCI, we found that no single variable consistently explained the CCI variation across the patient population. Each patient appeared sensitive to one or a few particular factors, but because of marked intraindividual variation, it was not possible to identify a priori which factors were important for a given patient. The single exception was a solid-phase red blood cell adherence assay used to cross-match platelets, but only for alloimmunized patients. We also evaluated the utility of requesting HLA-matched platelets from the local suppliers and maintained a clear distinction between platelets simply ordered as HLA matched and actually HLA-identical platelets. Accounting for the confounding clinical-, patient-, and blood bank-related factors, the cross-match assay was a better predictor of an adequate CCI than ordering platelets as HLA matched.

Effects of Pasteurella haemolytica A1 culture supernatant on mechanisms controlling bovine alveolar macrophage oxygen radical production.

Pasteurella haemolytica A1 culture supernatant containing leukotoxin, and modifiers of cyclic nucleotide and arachidonate metabolism, were evaluated for their ability to alter oxygen radical production by pulmonary alveolar macrophages obtained from seven Holstein calves. Calves were sedated, and underwent bronchoalveolar lavage to harvest macrophages, which were then incubated with culture supernatant and/or the drugs and toxins under study, and challenged with opsonized zymosan to induce oxygen radical generation. This was measured by a chemiluminescence technique. Pasteurella haemolytica A1 culture supernatant alone delayed the time to maximum oxygen radical production, although total production was increased. The cyclooxygenase inhibitor indomethacin and the phospholipase inhibitor p-bromophenacyl bromide significantly reduced maximum oxygen radical production, but their effects were diminished in the presence of culture supernatant. Although forskolin markedly inhibited oxygen radical generation, this effect was not altered by culture supernatant. Incubation of macrophages with pertussis toxin had no effect on oxygen radical production, while incubation with cholera toxin did inhibit production. This inhibitory effect was significantly lessened by concurrent incubation with P. haemolytica A1 culture supernatant.

Predictive velocity estimation in the pursuit reflex response to pseudo-random and step displacement stimuli in man.

1. Eye movements have been recorded in man in response to various forms of continuous and discontinuous target motions in the horizontal plane in an attempt to establish the mechanisms of prediction in the pursuit reflex. 2. In an initial experiment the target motion was composed of four sinusoids, each of peak velocity +/- 3.3 deg/s. The three lowest frequencies (0.11, 0.24 and 0.37 Hz) remained constant whereas the highest frequency (F4) was varied from 0.39 to 2.08 Hz. When F4 was 0.39 Hz, all frequency components had a high level of eye velocity gain (mean 0.92) but as F4 was increased there was a significant (P less than 0.001) decline in gain for all three low frequencies which reached a minimum (mean 0.53) when F4 was 1.56 Hz. However, the gain for F4 always remained at a high level, comparable to that evoked by a discrete frequency sinusoid of the same frequency. 3. When the highest-frequency sinusoid was replaced by a square wave of identical amplitude a similar decline in gain for the low frequencies was observed. Eye velocity exhibited a quasi-sinusoidal modulation at the frequency of the square wave even though the rapid steps did not constitute a suitable stimulus to the visual velocity feed-back mechanisms. 4. When only two sinusoids were mixed to form the pursuit stimulus a similar break-down in gain of the lower-frequency component was observed which reached a minimum (mean gain 0.58) when F2 was between 1 and 2 Hz. This implies that the predictability of stimulus motion is dependent, not on the complexity of the stimulus, but on its highest-frequency component. 5. Presentation of square-wave target displacements alone confirmed that smooth eye movements could be evoked by such a stimulus. Eye velocity was at a maximum between 1.0 and 1.5 Hz and was predictive of ensuing target displacement. Responses to staircase step sequences of varying duration indicated that prediction was based on an assessment of the duration of the preceding sequence. 6. Tachistoscopic presentation of targets during low-frequency sinusoidal oscillation indicated that illumination of the target for very short periods (10-320 ms) as few as two times per cycle during minimum velocity phases enhanced the perception of continuous movement. A predictive eye movement was evoked with a high level of peak velocity which then decayed until the subsequent exposure of the target.(ABSTRACT TRUNCATED AT 400 WORDS)