Perforin- and Fas-dependent mechanisms of natural killer cell-mediated rejection of incompatible bone marrow cell grafts

Natural killer (NK) cells eliminate target cells infected with intracellular pathogens and tumor cells by employing the granule exocytosis and death receptor pathways. They also mediate the acute rejection of incompatible bone marrow cell (BMC) grafts. However, the cytotoxic mechanisms employed during acute BMC graft rejection are obscure. Throughout these studies, BMC graft rejection was compared between two inbred strains of mice: 129 mice, which apparently use perforin- and Fas-dependent cytotoxicity, and C57BL/6 (B6) mice, which are able to exploit perforin- and/or Fas-independent mechanisms. Using perforin-knockout (PKO) mice, we have determined that the granule exocytosis pathway can play a major role in NK cell-mediated rejection of allogeneic and MHC class I-deficient BMC, depending upon the genetic background of the recipient and the environmental housing conditions. Although the granule exocytosis pathway seems to be the most potent cytolytic mechanism of NK cell-mediated rejection, alternative perforin-independent mechanisms, such as death receptor-induced apoptosis, also exist. By preventing both perforin- and Fas-mediated interactions concurrently, we observed that 129 mice were impaired in mediating MHC class I-deficient BMC rejection, while B6 mice maintained strong rejection capacities. The administration of neutralizing TNF antibodies to B6PKO mice before challenging with Fas and MHC class I double-deficient BMC still did not reverse rejection. Thus, our studies reveal the relative importance of perforin-, Fas-, and TNF-based cytotoxicity in NK cell-mediated rejection of incompatible BMC.     

Susceptibility of corneal allografts and xenografts to antibody-mediated rejection

The effects of passive transfer of antisera containing cytotoxic antibodies to allo- and xenoantigens on survival of corneal allografts and xenografts were evaluated in experimental models. Corneas from allogeneic B10 or xenogeneic rat Lewis donors were grafted orthotopically into BALB/c mice. Recipient mice were treated with donor-specific antisera administered at the period of grafting or at 2 weeks after transplantation. Rejection was determined by the severity of corneal opacity using a standard scoring system. Treatment of graft recipients with donor-specific antisera accelerated the onset of graft rejection and significantly shortened survival times of both corneal allografts and xenografts. Corneal xenografts, which had been accepted after treatment with anti-CD4 monoclonal antibody, were acutely rejected by the passive transfer of antiserum against xenoantigens. The results suggest that corneal grafts are vulnerable to antibody-dependent immunity and that cytotoxic antibodies against graft donor antigens can mediate rejection of both corneal allografts and xenografts.     

A Field-Compatible Method for Interpolating Biopotentials

Mapping of bioelectric potentials over a given surface (e.g., the torso surface, the scalp) often requires interpolation of potentials into regions of missing data. Existing interpolation methods introduce significant errors when interpolating into large regions of high potential gradients, due mostly to their incompatibility with the properties of the three-dimensional (3D) potential field. In this paper, an interpolation method, inverse-forward (IF) interpolation, was developed to be consistent with Laplace's equation that governs the 3D field in the volume conductor bounded by the mapped surface. This method is evaluated in an experimental heart–torso preparation in the context of electrocardiographic body surface potential mapping. Results demonstrate that IF interpolation is able to recreate major potential features such as a potential minimum and high potential gradients within a large region of missing data. Other commonly used interpolation methods failed to reconstruct major potential features or preserve high potential gradients. An example of IF interpolation with patient data is provided to illustrate its applicability in the actual clinical setting. Application of IF interpolation in the context of noninvasive reconstruction of epicardial potentials (the inverse problem) is also examined. © 1998 Biomedical Engineering Society. PAC98: 8710+e, 0260Ed     

A Saccharomyces cerevisiae mutant defective in the kinesin-like protein Kar3 is sensitive to NaCl-stress

Several mutants of Saccharomyces cerevisiae showing poor growth in the presence of elevated concentrations of NaCl were isolated to identify genes involved in the osmo-stress response. One of these mutants (WAY.5-4A-11; osr11) which showed a clear 2:2 segregation of the salt-stress phenotype upon tetrad analysis when crossed to a wild-type strain has been characterised. The mutation responsible for poor growth under salt-stress was recessive. The corresponding gene was cloned by complementation of the mutant phenotype and a 3.5-kb fragment was isolated. The sequence of this fragment matched that of KAR3, a gene previously identified to be involved in karyogamy and mitosis. Allelism of OSR11 to KAR3 was confirmed by tetrad analysis, and disruption mutants showed the same NaCl-phenotype as the original osr11 mutation. The disruption mutant was more sensitive to high sucrose concentrations than the original mutant was to high glucose concentrations. In a different genetic background (W303-1A), the kar3 disruptants were less sensitive to osmo-stress than the WAY.5-4A strain. Heat-stress, nitrogen-starvation and cultivation on ethanol failed to affect the growth of osr11 and kar3 mutants, pointing to a possible specific involvement of KAR3 in the osmotic-stress response. Microscopic studies showed that cell division of the kar3 mutants was impaired and NaCl-stress conditions aggravated the phenotype. Received: 7 April / 21 July 1997     

Psychometric properties of the SUNYA revision of the psychosomatic symptom checklist

The Psychosomatic Symptom Checklist (PSC), a questionnaire assessing psychosomatic symptoms, was administered to two separate samples of college students. For Sample 1 (N=698),the questionnaire was readministered to three separate subsets at intervals of either 1 week (N=143),4 weeks (N=74),or 8 weeks (N=48).Each subset of subjects recompleted the PSC on only one of the three retest intervals. Based on the initial administration an analysis of the normative data revealed a mean total score of 23.7, suggesting a relatively low degree of psychosomatic symptoms in this group. Although total scores decreased slightly over time, test-retest correlations remained high (r>0.80, P<0.0001).Individual item correlations varied and also decreased across time; however, the majority of correlations was greater than r=0.50 throughout. Sample 2 (N=249)completed the PSC, Beck Depression Inventory (BDI), State-Trait Anxiety Inventory (STAI-X), and Rathus Assertiveness Scale (RAS), and intercorrelations were computed between these measures. This analysis revealed little overlap between the psychosomatic complaints assessed by the PSC and other commonly used measures of psychological distress. Finally, a factor analysis revealed one major factor on which all but 2 of the 17 questionnaire items loaded significantly. These results suggest that the PSC is sensitive to psychosomatic distress and remains reliable over time.This reaserch was supported in part by Grants NS-15235 and NS-16891 from NINCDS.     

Cytogenetic diagnosis using midtrimester fetal blood samples: Application to suspected mosaicism and other diagnostic problems

Cytogenetic studies on fetal blood cells obtained at 18–25 weeks gestation have provided information for decision making in 25 cases identified as being at high risk of having an abnormal fetus. In particular, in the 21 cases studied to consider the possibility of true mosaicism, confirmation in fetal blood was obtained in three, one of which presented as a pseudomosaic on the original amniotic fluid cell study. Fetal blood was also informative in two cases (one positive and the other negative) in which a diagnosis of the fragile X syndrome was being considered. Furthermore, when high risk pregnancies presented late in gestation (21–24 weeks), these methods allowed for a rapid cytogenetic diagnosis. The procedure has proved useful in most of these cases since the couples involved had indicated that they would probably have terminated the pregnancy without the reassurance of normal fetal lymphocyte studies. Since the technique carries a much higher risk of pregnancy loss than does amniocentesis, its use should only be considered when there are compelling indications.     

Genetic mapping ofPim-1 putative oncogene to mouse chromosome 17

Pim-1 is a putative oncogene activated in T-cell lymphomas induced by Moloney and AKR mink cell focus forming (MCF) viruses. We have determined the chromosomal localization of the Pim-1gene in mice by Southern blot analysis of DNAs obtained from a panel of mouse-Chinese hamster somatic cell hybrids. The Pim-1gene was localized on chromosome 17, a chromosome frequently aberrant in T-cell lymphomas. Two chromosomal regions, containing sequences homologous to regions within the Pim-1locus, were localized on chromosome 6 and 16.     

Identification of two point mutations and a one base deletion in exon 19 of the dystrophin gene by heteroduplex formation

Two thirds of the Duchenne muscular dystrophy population haveeither gene deletions or duplications. The nondeletion/duplicationcases are most likely the result of point mutations or smalldeletions and duplications that cannot be easily identifiedby current strategies. The major obstacle in identifying smallmutations is due to the large size of the dystrophin gene. Weselectively screened 5 DMD exons containing CpG dinucleotidesin 110 DMD patients without detectable deletions or duplications.Nonsenses mutations are frequently due to a C- to -T transitionwithin a CG dinucleotide pair. To screen for the nonsense mutations,we used the heteroduplex method. Utilizing this approach, weidentified 2 different nonsense mutations and a single basedeletion all occurring in exon 19. This is the first reportof a clustering of small mutations in the the dystrophin gene.