Extreme ultraviolet light from a tin plasma driven by a 2-µm-wavelength laser.

An experimental study of laser-produced plasmas is performed by irradiating a planar tin target by laser pulses, of 4.8 ns duration, produced from a KTP-based 2-µm-wavelength master oscillator power amplifier. Comparative spectroscopic investigations are performed for plasmas driven by 1-µm- and 2-µm-wavelength pulsed lasers, over a wide range of laser intensities spanning 0.5 - 5 × 1011 W/cm 2. Similar extreme ultraviolet (EUV) spectra in the 5.5-25.5 nm wavelength range and underlying plasma ionicities are obtained when the intensity ratio is kept fixed at I1µm/I2µm = 2.4(7). Crucially, the conversion efficiency (CE) of 2-µm-laser energy into radiation within a 2% bandwidth centered at 13.5 nm relevant for industrial applications is found to be a factor of two larger, at a 60 degree observation angle, than in the case of the denser 1-µm-laser-driven plasma. Our findings regarding the scaling of the optimum laser intensity for efficient EUV generation and CE with drive laser wavelength are extended to other laser wavelengths using available literature data.

Prominent radiative contributions from multiply-excited states in laser-produced tin plasma for nanolithography.

Extreme ultraviolet (EUV) lithography is currently entering high-volume manufacturing to enable the continued miniaturization of semiconductor devices. The required EUV light, at 13.5 nm wavelength, is produced in a hot and dense laser-driven tin plasma. The atomic origins of this light are demonstrably poorly understood. Here we calculate detailed tin opacity spectra using the Los Alamos atomic physics suite ATOMIC and validate these calculations with experimental comparisons. Our key finding is that EUV light largely originates from transitions between multiply-excited states, and not from the singly-excited states decaying to the ground state as is the current paradigm. Moreover, we find that transitions between these multiply-excited states also contribute in the same narrow window around 13.5 nm as those originating from singly-excited states, and this striking property holds over a wide range of charge states. We thus reveal the doubly magic behavior of tin and the origins of the EUV light.

Bracing for survival.

As health care continues to move toward smaller, more consolidated support services, materiel managers will face constricted promotion and employment opportunities. Flexibility and a willingness to embrace change as the occasion for personal and professional advancement will be two hallmarks of success. This article focuses on outsourcing and integrated delivery networks by discussing the challenges and opportunities they present.

Evidence that the antigen receptors of cytotoxic T lymphocytes interact with a common recognition pattern on the H-2Kb molecule.

Recognition of class I MHC antigens involves interaction between TCRs of cytotoxic T lymphocytes (CTL) and the two alpha helices of MHC molecules. Using a combined panel of H-2Kb mutants selected by either a CTL clone or MAbs, we have shown evidence that the TCRs of 59 Kb-specific CTL clones shared a common binding pattern on the H-2Kb molecule. Mutations of amino acid residues at the C-terminal regions, but not the N-terminal regions, of the alpha helices abrogated the recognition by the majority of the clones. The data suggests that TCRs predominantly recognize the class I MHC molecule with an orientation that is parallel to the beta-pleated strands and diagonal to the alpha helices.

Superantigens and their role in infectious disease.

Although the exact mechanisms by which superantigens may contribute to the pathogenesis of diseases are unknown, it seems increasingly likely that they have a role in the induction and pathogenesis of disease. The studies described here demonstrate that in several different diseases either bacterial or viral superantigens can be isolated from patients. There is also a preferential expansion of particular V beta T-cell subsets, which is a common feature of superantigen stimulation. From the work that has been done to date it can be hypothesized that superantigens may act in several ways. They may stimulate and activate T cells that are autoreactive and lead to the induction or exacerbation of autoimmune disease, as in RA. Alternatively, they may lead to the depletion of T-cell subsets based on V beta expression, thereby resulting in the severe reduction in lymphocytes in certain immunodeficiency diseases such as AIDS. But perhaps the most likely contribution of superantigens to disease pathogenesis is seen indirectly by their effect on the immune system-particularly the stimulation of large numbers of T lymphocytes expressing the same V beta domain. Thus it is likely that the direct effect of various T-cell-derived inflammatory mediators (i.e., interleukins and other cytokines) released by these activated T lymphocytes is the primary cause of disease pathology via response to superantigen stimulation. In addition to the diseases discussed here, there are a number of other diseases in which a potential role for superantigens is being studied. These include autoimmune diseases seen after group A streptococcal infections in which the streptococcal M protein has been postulated to act as a superantigen such as scarlet fever, rheumatic heart disease, and poststreptococcal glomerulonephritis. Other diseases being studied include psoriasis, lupus-like disease, and lymphoproliferative diseases (reviewed in Kotzin et al.). In the coming years the exact role of superantigens and the specific mechanisms by which they contribute to disease should be more clearly defined. Our understanding of these molecules could also lead to new therapies for the treatment of these diseases.

Presentation of a horse cytochrome c peptide by multiple H-2b class I major histocompatibility complex (MHC) molecules to C57BL/6- and bm1-derived cytotoxic T lymphocytes: presence of a single MHC anchor residue may confer efficient peptide-specific CTL recognition.

In this study the immunogenic tryptic fragment from a horse cytochrome c (cyt c) digest recognized by cytotoxic T lymphocytes (CTL) induced by in vitro peptide stimulation from C57BL/6 (B6) and mutant B6.C-H-2bm1 (bm1) mice is identified. An identical sequence, p40-53, is recognized by CTL from both B6 and bm1 mice. In addition, both B6 bm1 cloned CTL lines display unusual major histocompatibility complex (MHC) class I-restricted recognition of this peptide in that they respond to it in the context of H-2Kb, H-2Db, and H-2Kbm1 class I molecules, although the sequence lacks the usual structural Kb and Db peptide-binding motifs. Truncated analogues which resemble the lengths of naturally processed MHC class I-presented peptides, confer reactivity for B6 and bm1 CTL against EL4 (H-2b) targets as well as the L cell transfectants, L+Kb, L+Db, and L+Kbm1. The antigenic peptide with the greatest potency is p41-49, which appears to be generated by angiotensin converting enzyme cleavage of the full-length p40-53 tryptic peptide. The minimum antigenic peptide recognized by both B6 and bm1 CTL, and which targets lysis on each of the transfectants, is the hexamer p43-48 peptide from horse cyt c. Residues Pro44 and Thr47, which occupy polymorphic positions with respect to other species-variant cyt c molecules, influence recognition of these peptides differently for the B6 and bm1 CTL. The ability of H-2Kb, H-2Db, and mutant H-2Kbm1 class I molecules to present the same peptide to a single cloned CTL is discussed in the context of current knowledge of peptide anchor residues and side chain-specific binding pockets in the MHC class I peptide-binding site.

Minor pocket B influences peptide binding, peptide presentation and alloantigenicity of H-2Kb.

Microsequence analysis of peptides eluted from the murine class I H-2Kb molecule together with the three-dimensional structure of the molecule co-crystallized with a homogeneous population of peptides suggests that pocket B is a minor pocket that does not play a major role in peptide presentation. This is in contrast to most other class I molecules in which pocket B plays a central role in selecting and presenting antigenic peptides. To investigate the role of pocket B in antigen presentation by the Kb molecule, we analyzed site-directed mutants of position 45 in pocket B for their effect on both allo- and peptide-specific recognition. We made an identical set of mutations in Kbm8 at residue 45 in order to evaluate their influence in the context of a more open pocket B which results from the bm8 substitution at amino acid 24 (E-->S). We demonstrated that this minor pocket did play a significant role in the antigenicity of both molecules and that this role was more readily apparent in the context of the more open pocket B of Kbm8. In addition, we found that some substitutions of residue 45 in the Kbm8 molecule restored recognition by some alloreactive and peptide specific anti-Kb T cell clones which are normally restricted to Kb, indicating that multiple configurations of amino acids in a pocket could result in similar binding and presentation capabilities.

Selective reactivity of CD8-independent T lymphocytes to a cytotoxic T lymphocyte-selected H-2Kb mutant altered at position 222 in the alpha 3 domain.

To study the structural basis for specificity and affinity of cytotoxic T lymphocytes for major histocompatibility complex/peptide complexes, we have employed a cytotoxic T lymphocyte (CTL)-mediated immunoselection approach to obtain H-2Kb structural mutants which are resistant to lysis by a Kb-specific alloreactive CTL clone. In this study we describe the Kb structural mutant, designated R8.60.14, recovered following immunoselection using the CD8-dependent CTL clone 60 as a selective agent. Although serologically unaltered with respect to Kb expression, R8.60.14 is not recognized by CD8-dependent, Kb-specific CTL. DNA sequence analysis revealed a single Glu----Lys amino acid substitution at position 222 in the Kb alpha 3-domain of this variant. To determine if a direct correlation exists between CD8 dependence of a Kb specific CTL and its failure to respond to R8.60.14, we examined the lytic response against R8.60.14 by CD8-independent, Kb-specific CTL obtained from long-term culture in the presence of anti-CD8 monoclonal antibody, 3.155. CD8-independent CTL exhibit no difference in their response against the R8 parent and R8.60.14 variant. This study demonstrates unequivocally that Kb-specific recognition of R8.60.14 by CD8-independent CTL is unaltered, while the response by CD8-dependent CTL is completely abrogated. Thus, the sole basis for emergence of this variant in the CTL-mediated immunoselection approach used in this study resides in the alteration of a single CD8-binding site residue at position 222 in the Kb alpha 3 domain. The functional importance of this Glu222 residue for the interaction between the CD8 molecule on CD8-dependent CTL and the Kb alpha 3 domain is further reinforced by virtue of the recovery of the R8.60.14 variant on the basis of its resistance to lysis by a CD8-dependent CTL clone in this CTL-mediated immunoselection approach.

Identification of an autologous insulin B chain peptide as a target antigen for H-2Kb-restricted cytotoxic T lymphocytes.

We have examined the CD8+ peripheral T cell repertoire of C57BL/6 (H-2b) mice for cytotoxic T lymphocyte (CTL) reactivities to insulin, using in vitro immunization with a chymotryptic digest of reduced bovine insulin. The results presented in this study demonstrate that potentially autoreactive H-2Kb-restricted cytotoxic T cells specific for an autologous insulin B chain peptide are present in the preimmune splenic T cell repertoire. The immunogenic peptide comprises residues 7-15 from the insulin B chain and has features in common with naturally processed Kb-restricted peptides identified by others. The minimal peptide sequence recognized by these cytotoxic T cells is 10-15, which is highly conserved in mammalian species and constitutes a self-peptide in mice. The presence of class I major histocompatibility complex-restricted CTLs with potentially autoreactive specificities in preimmune animals raises the possibility of a role for such cells in autoimmune disease states. Possible mechanisms for the in vivo expansion of insulin peptide-specific CTLs are discussed.

Delivery following cesarean section and perinatal mortality.

The perinatal mortality in 1498 patients with one or more previous cesarean section scars delivered between 1972 and 1982 was analyzed. Repeat elective cesarean section was performed in 654 (44%) patients and 844 (56%) were subjected to a "trial of scar." Successful vaginal delivery occurred in 702 (83%) patients and 142 (17%) had emergency repeat operations. There were 46 perinatal deaths, giving a perinatal mortality rate (PMR) of 30.3/1000. It was lowest in patients who electively had a cesarean section (10.6/1000). The corrected PMR was twice as high in the trial of scar group. The PMR for the overall hospital population (27,072 babies) during the study period was 22.5/1000. There were four perinatal deaths in association with dehiscence of the uterine scar. The PMR in trial of scar patients decreased from 40/1000 to less than 20/1000 without a major change in policy. Meanwhile the unit cesarean section rate increased from 5 to 10%, but the repeat section rate was consistent at around 38.5%. Regional analgesia was used in 192 patients, for repeat section in five and trial of scar in 187. Oxytocin was given to 546 (65%) patients. Scar rupture is considered the major contraindication to a trial of scar, but emphasis was not laid on the possible increased perinatal mortality with this procedure before the 1980s. In view of the improvement in the PMR and the added risk to the mother with cesarean delivery, we advocate a policy of trial of scar with informed consent in selected cases.

Use of mutants to analyze regions on the H-2Kb molecule for interaction with immune receptors.

MHC variants isolated both in vivo (by tissue graft rejection) and in vitro (by antibody selection) were utilized to study sites on the H-2Kb molecule involved in interaction with antibodies and with the TCR. Kb mutants selected by antibodies were found to have single point mutations, which when analyzed in the context of the three-dimensional structure of a Kb molecule modeled from HLA-A2 coordinates showed that the altered residues were localized mostly to the alpha 1 and alpha 2 helices. The side chains of the variant amino acid residues pointed upward and away from the antigenic site. Analysis of the altered amino acids in the previously described tissue-graft-selected Kb mutants showed that the side chains of the variant residues occur either in the alpha-helical regions or in the beta-pleated-sheet floor of the antigen groove, but every mutant contained at least one and sometimes several acid side chains projecting into the antigen-binding groove. Monoclonal antibody studies showed that the available monoclonal antibodies mapped to discrete domain-specific sites. Analysis of CTL recognition sites using cloned mutant anti-parent or allogeneic combinations showed that all CTL clones examined interacted with amino acid side-chain residues on both the alpha 1 and alpha 2 helices. Thus, we concluded that the CTLs must simultaneously interact with the amino acid residues in both the alpha-helical stretches of the alpha 1 and alpha 2 domains. Our analyses with the point mutants imply that the TCR must interact with the MHC molecule over a relatively large surface area in such an orientation that it interfaces with the two alpha helices, as well as with the foreign or self-peptide in the antigen-binding site between the helices. These findings, together with the observation that several of the in vivo Kb mutants induce strong alloreactions yet have changes only in the bottom of the antigen-binding groove and no alterations in the alpha-helical residues, are consistent with the hypothesis that in some cases alloreaction can be the result of T-cell recognition of an altered pattern on the MHC molecule due to a changed peptide in the antigen groove.

Induction of cytotoxic T lymphocytes by primary in vitro stimulation with peptides.

Antigen-specific cytotoxic T cells can be generated by primary in vitro stimulation of spleen cells from C57BL/6 mice with appropriate peptide fragments. This response can be elicited without prior in vivo immunization. Chicken OVA fragmented with either cyanogen bromide (CN OVA) or trypsin (T OVA) was used as a source of mixed peptides. A synthetic peptide, NP365-380, representing the sequence 365-380 from influenza virus A/PR/8 nucleoprotein, was also used, since this contains the main determinants recognized by CTL generated from H-2b mice infected with A/PR/8 virus. The primary in vitro cytotoxic T cell response was peptide specific, since targets were lysed only in the presence of appropriate peptide antigens. Native OVA could not elicit primary effectors in vitro nor could it sensitize targets for lysis by OVA digest-specific CTL. A synthetic peptide corresponding to residues 111-122 within the OVA sequence could sensitize targets for lysis by effectors induced against T OVA. Effectors generated by in vitro stimulation were CD8+, CD4-, and H-2Db-restricted for NP365-380 and T OVA recognition. CN OVA-specific effectors were also CD8+, CD4-, but surprisingly, were able to lyse a range of H-2-different targets in an antigen-specific manner. These effectors failed to lyse a tumor line that does not express class I MHC molecules. This broad MHC restriction pattern was also apparent at the clonal level. In all cases, the antipeptide CTL generated by primary in vitro stimulation were inefficient in lysing target cells expressing endogenous forms of antigens, such as influenza virus-infected cells or cells transfected with the OVA cDNA. However, cytotoxic T cell lines generated in vitro against the NP365-380 peptide did contain a minor population of virus-reactive cells that could be selectively expanded by stimulation with A/PR/8-infected spleen cells. These results are discussed in terms of class I-restricted T cell stimulation in the absence of antigen processing by high surface densities of peptide/MHC complexes.

Characterization of dual-reactive H-2Kb-restricted anti-vesicular stomatitus virus and alloreactive cytotoxic T cells.

Cross-reactive recognition of alloantigen by "self + X"-reactive cytotoxic T lymphocytes (CTL) has been documented in a variety of systems. It has been shown previously that the H-2Kb-restricted CTL response of C57BL/6 (B6) mice to vesicular stomatitis virus (VSV) infection is partially cross-reactive on uninfected target cells expressing the H-2Kbm8 mutation. In this report, we describe the isolation and detailed characterization of such dual-reactive CTL. By employing EL4 tumor lines transfected with genes encoding various VSV proteins, we demonstrated that the majority of dual-reactive CTL recognize the internal N protein of VSV and are also reactive against uninfected bm8 targets. Although the response of normal B6 mice to bm8 stimulators shows no measurable cross-reactivity on VSV-infected targets, the response of VSV-primed B6 mice to bm8 stimulation is almost entirely cross-reactive, lysing VSV-B6 targets and uninfected bm8 targets roughly equally. Furthermore, about 70% of CTL clones isolated from such mice by bm8 stimulation are dual-reactive with respect to effector function. Analysis at the population and clonal levels with cold target competition and antibody blocking suggests that the bulk of dual-reactive CTL have a higher avidity for VSV-B6 targets than for bm8 targets. The extreme case of this is illustrated by a fraction of CTL clones, isolated and maintained on bm8 stimulators, which lyse VSV-B6 targets but do not lyse bm8 targets. One such CTL clone is shown to be specific for the bm8 antigen in proliferation assays. These results demonstrate that: the specificity of an alloreactive CTL response may be dramatically altered by previous antigenic encounters; and dual-reactive CTL display a significant difference in affinity of the CTL receptor-determinant interaction, depending on the target which is recognized.

Immunoselection of structural H-2Kb variants: use of cloned cytolytic T cells to select for loss of a CTL-defined allodeterminant.

Functional studies concerning the unique interaction between class I H-2 allodeterminants and cytolytic T lymphocyte (CTL) antigen receptors have benefitted from the development of H-2Kb mutant mouse strains and somatic H-2 variants selected with monoclonal antibody. Here, we describe the development of a novel approach to immunoselection of somatic H-2Kb variants employing a Kb-specific CTL clone as the negative selective agent. The rationale for this method is that the use of an alloreactive CTL clone as the selective agent should enable us to detect the emergence of structural Kb variants based on their loss of the relevant CTL-defined allodeterminant. Thus, these structural variants are well suited to an in-depth analysis of the functional relationship between H-2 antigens and receptor recognition by CTL. Using this approach, we successfully isolated two types of structural Kb variants, as well as numerous Kb-loss variants. The functional studies described in this paper indicate that these structural variants exhibit alterations in expression of both CTL-defined and serologically defined H-2Kb allodeterminants. The structural characterization of such variants should enable us to identify the precise amino acid residues responsible for the creation of the relevant CTL-defined Kb allodeterminants.

Susceptibility of Adenovirus 2-transformed rat cell lines to natural killer (NK) cells: direct correlation between NK resistance and in vivo tumorigenesis.

The susceptibility to natural killer (NK)-mediated cell lysis of Adenovirus type 2 (Ad2)-transformed rat embryo fibroblast cell lines, which differed markedly in tumorigenic potential in vivo (T2C4 greater than F19 greater than F17), was investigated by using NK effector cells from F344 rat or athymic nude rat spleens. A comparison of the degree of NK-mediated lysis obtained with these tumor cell targets suggested a direct relationship between the resistance of a cell to NK cell lysis and its potential to form tumors in vivo. The cells were lysed in the following order of increasing susceptibility: T2C4 less than F4 less than F19 less than F17. Whether T cells or macrophages played a significant role in the observed lytic activity was determined by treating the NK effector cell population with anti-rat T cell serum (alpha T) and complement or by depletion of macrophages after binding to a glass bead column and treatment with carbonyl iron. A series of clonal sublines derived from the parental F17 and F4 cell lines further strengthened this relationship between tumorigenesis and resistance to NK-mediated cell lysis. Tumorigenic subclones from the non-tumorigenic F17 parental cells were demonstrated to be comparatively resistant to NK-mediated lysis. Tumorigenic subclones from tumorigenic F4 parental cell population showed a susceptibility to NK-mediated cell lysis virtually identical to the parental F4 cells. The implication of these results are discussed.

The effect of danazol on the production of C1 inhibitor in the guinea pig.

The production of C1 inhibitor (C1-INH) in guinea pig liver was studied following administration of danazol. Immunochemical assays of the inhibitor were performed on serial serum specimens. An increase in serum C1-INH was observed following danazol treatment. Immunohistochemical studies confirmed that the liver is the sole site of synthesis of C1-INH in guinea pigs. This appears to occur in certain clones of hepatocytes. Ultrastructural studies revealed an increase in coarse endoplasmic reticulum and mitochondria in danazol-treated animals. In in vitro studies with liver slices, however, more C1-INH was detected in culture supernatants from untreated animals than from danazol-treated animals. This observation suggests that secretion of C1-INH from stimulated hepatocytes may require additional factors which are present in vivo but not in vitro.

Intestinal lactase, sucrase and alkaline phosphatase in relation to age, sex and site of intestinal biopsy in 477 Irish subjects.

Small intestinal lactase, sucrase and alkaline phosphatase activities were measured in histologically normal peroral intestinal biopsies from 477 individuals. Enzyme activities varied with age, sex, site of biopsy, and were lowest in post-weaning children and highest in young adults. Lactase activity does not decrease with advancing age.

Biologic properties of bacterial lipopolysaccharides treated with chromium chloride.

Addition of small amounts of chromium chloride to a saline suspension of Salmonella typhosa lipopolysaccharide (LPD; Difco) caused a marked reduction in several of the biologic activities of this substance including toxicity, B-cell mitogenicity, plasma colony-stimulating activity (CSA), radioprotective effect, and induction of the dermal Shwartzman reaction. Nevertheless, LPS treated with chromium chloride was found to be at least as effective as untreated LPS in enhancing resistance of B6CBF1 mice to the lethal effects of Klebsiella pneumoniae infection.