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D S Strayer

Publications and source records attributed to D S Strayer.

At least 19 recordsLinked to original sources
PubMed1995

DNA binding proteins that amplify surfactant protein B gene expression: isolation and characterization.

We identified and characterized two proteins that bind the promoter of surfactant protein B (SP-B) and affect its expression. Proteins A2 and B were identified and their cDNAs cloned and sequenced. Both were novel. They bound a 212-bp functional promoter region at an NF1 site, located between -184 and -198. Effects of these DNAbp on SP-B promoter activity were studied by contransfecting a reporter construct of this 212-bp sequence + luciferase, together with expression constructs for A2 and B into H441 cells. Alone, A2 and B expression elicited modest but statistically significant increases in SP-B promoter activity. When dexamethasone was added, B further increased SP-B promoter activity. For SP-B, basal expression and glucocorticoid responsiveness may involve a number of hitherto unknown gene activators.

Adult↗
PubMed1995

Levels of SP-A-anti-SP-A immune complexes in neonatal respiratory distress syndrome correlate with subsequent development of bronchopulmonary dysplasia.

As part of a double-blind, randomized, placebo-controlled study of human surfactant therapy for neonatal respiratory distress syndrome (NRDS), we measured circulating immune complexes between surfactant protein-A and anti-surfactant protein-A antibodies (SAS). Plasma from almost all infants contained detectable immune complexes. Immune complex levels in surfactant-treated infants were comparable with those of placebo-treated controls. Despite the relatively small sample size, maximum SAS immune complex values between 2 and 4 weeks after birth correlated significantly with subsequent development of BPD. Levels of these immune complexes correlated with eventual BPD independently of, and more strongly than, gestational age and birth weight. Thus, plasma SAS immune complex measurements may be useful in analyzing the course and outcome of NRDS, in particular the likelihood of subsequent development of BPD. This assay may also help to identify infants at risk for BPD and to target preventative therapy to them.

Antigen-Antibody Complex↗
PubMed1995

Glucocorticoid responsiveness conferred by a cloned DNA binding protein.

Glucocorticoids stimulate surfactant protein-B (SP-B) (expression in type II alveolar cells) by unknown mechanisms. We identified, cloned, and characterized a protein that binds the SP-B promoter. This protein, D, increases the activity of the SP-B promoter in response to glucocorticoid stimulation. Protein D was identified by its ability to bind the SP-B promoter region, which it binds at an NF1 site from -184 to -198 bp. Its binding was abolished by digestion of promoter DNA with BalI, which cuts at -194. Protein D was cloned and sequenced. It is a new DNA binding protein of 33 kDa whose carboxyl end contains a modified basic leucine zipper-like DNA binding motif (bzip). The effects of D on SP-B promoter activity were studied in H441 cells, using a reporter construct containing 212 bp from the SP-B promoter with a luciferase reporter gene (p2121uc), which was cotransfected with a protein D expression construct in which D expression was controlled by the SV40 early promoter. These two plasmids were cotransfected into H441 cells. Without added glucocorticoids, D did not alter SP-B promoter activity. When dexamethasone was added, D strongly enhanced SP-B promoter activity. Identification of this protein suggests that, at least for SP-B, glucocorticoid responsiveness may involve one or more hitherto unknown gene activators.

Amino Acid Sequence↗
PubMed1993

Effect of virus infection on levels of transcripts for cellular genes.

Malignant rabbit fibroma virus (MV) induces tumors composed of proliferating cells, principally fibroblasts, and vasculature. These tumors are associated with large amounts of collagen and other connective tissue proteins. We studied the effect of MV infection on levels of mRNA for alpha 1 chains of collagens type I, III and V in RK-13 fibroblasts and alpha 2 chain of collagen type I. MV infection induces expression of specific collagen genes at particular time points after infection in vitro. Expression of these collagen genes is clearly different in MV-infected cells compared to uninfected cells. Transcript levels for a cellular transcription factor that regulates expression of alpha 1 chain type I collagen, cbf-a, were increased in MV-infected cells prior to the increase in type I collagen mRNA. The virus infection also specifically induced increased levels of mRNA for the cellular transcription factors c-fos and SP1. MV infection is therefore associated with increased levels of specific cellular mRNAs, and is correspondingly associated with increased mRNA for transcription factors that may regulate transcription of these genes. The ability of malignant fibroma virus to influence expression of cellular genes may be exerted through the cells' own transcription regulatory apparatus.

Animals↗
PubMed1992

Sequence and analysis of the BamHI "D" fragment of Shope fibroma virus: comparison with similar regions of related poxviruses.

Differences observed in the virulence of two related leporipoxviruses are closely tied to a particular region of their genomes. For the virulent poxvirus of this pair, malignant rabbit fibroma virus (MV), this region is the BamHI "C" fragment, which is 10.7 kb. For the avirulent poxvirus, Shope fibroma virus, SFV, this region is the corresponding BamHI "D" fragment, which is 13.1 kb. As part of our attempt to understand the virulence of these two viruses, we sequenced these two DNA fragments. The sequence for the BamHI "C" fragment of MV is reported elsewhere (Strayer et al., 1991). We report here the sequence for SFV's BamHI "D" fragment and resultant open reading frames, and compare both DNA and open reading frame structures to those of MV and other known poxviruses. The BamHI "D" fragment of SFV contains 12 open reading frames of 100 amino acids or more, arranged similarly to orf's in MV and vaccinia. Striking similarities between SFV and MV are seen in certain parts of this restriction fragment, including substantial stretches of DNA in which the two viruses are identical. Clear homologies exist between these leporipox virus genomes and those of other related poxviruses. To understand the pathogenesis of virus infection, one must appreciate the structure of those viral genes that play important roles in infection.

Base Sequence↗
PubMed1992

Applications of anti-curosurf antibodies to understanding the biology of alveolar surfactant.

The structure of surfactant-binding proteins from alveolar cell membranes was investigated using monoclonal anti-idiotypic antibodies directed against surfactant-binding regions of anti-surfactant monoclonal antibodies. Two different series of monoclonal anti-surfactant antibodies were used. One series included antibodies against rabbit surfactant, and the other antibodies against porcine surfactant. In addition, two monoclonal anti-idiotype antibodies were made. These anti-idiotype antibodies, called A2R and A2C, bind both anti-Curosurf and anti-rabbit surfactant antibodies comparably. They recognize a 30-kDa cell membrane protein on alveolar and bronchial epithelial cells. In addition, A2C and A2R block the binding of radiolabeled surfactant to these cells. Using a combination of A2C and A2R cDNA expression libraries from both human and porcine lungs were screened. One independent clone was identified in each library that produced protein that bound these monoclonal antibodies. The protein encoded by the cDNA in each clone bound radiolabeled recombinant surfactant protein-A. It is concluded that human and porcine alveolar cell SP-A binding proteins have been identified and its cDNA cloned. The potential implications of this finding for the understanding and treatment of surfactant deficiency states are considerable.

Animals↗
PubMed1991

Sequence and analysis of a portion of the genomes of Shope fibroma virus and malignant rabbit fibroma virus that is important for viral replication in lymphocytes.

The 10.7-kb BamHI "C" restriction fragment of malignant rabbit fibroma virus (MV) contains genes that are important for its immunosuppressive activity. When this fragment is transferred to a related avirulent leporipoxvirus, Shope fibroma virus (SFV), recombinant viruses show clinical features characteristic of MV: they replicate in lymphocytes and alter immune function in vitro, induce disseminated tumors in recipient rabbits, and are immunosuppressive in vivo. The 10.7-kb BamHI "C" restriction fragment of MV was sequenced in its entirety. Its DNA sequence and the 14 ORF's derived from analyzing this sequence are discussed. Analysis of known open reading frames to which the ORF's from MV's Bam "C" fragment show homology permits us to identify some MV ORF's showing high degrees of similarity to known and postulated proteins produced by vaccinia virus. Functions for some of these vaccinia proteins are known, while functions for others are hypothetical or unknown. Further analysis of genetic determinants of MV's virulence has indicated that two overlapping restriction subfragments of the BamHI "C" fragment can transfer MV's virulent behavior to SFV. The 0.7-kb region in which these two subfragments overlap includes the C-terminus of MV orf C-7 and the N terminus of MV orf C-8. These correspond to the C- and N-termini, respectively, of SFV orf's D-9 and D-10 and to vaccinia orf's D-6 (early transcription factor) and D-7 (subunit of RNA polymerase). We sequenced the region of SFV's BamHI "D" fragment in this area and illustrate here the comparative sequences of this portion of SFV's genome and orf's. On the basis of comparisons between MV, SFV, and vaccinia in this area we discuss the potential significance of these observations.

Amino Acid Sequence↗
PubMed1991

Immunogenicity of surfactant. I. Human alveolar surfactant.

The immunogenicity of lung surfactant derived from amniotic fluid has been well established. We have set out to examine the antigenic similarity of human surfactant to non-human alveolar surfactants currently being used therapeutically in clinical trials with neonatal respiratory distress syndrome. To this end, we raised a series of eight monoclonal antibodies in rats directed to human surfactant (H1 to H8). All antibodies bound human surfactant as measured by ELISA. Four of these monoclonal antibodies bound surfactant components by Western blot analysis: all bound a 9-10-kD species. In addition, one antibody (H2) bound a protein of 16 kD, one (H8) a 6-kD protein, and one (H6) a 30-kD protein. When mixed with surfactant, three antibodies, H4, H7 and H8, profoundly altered surfactant activity in vitro in the pulsating bubble surfactometer. Three other antibodies, H1, H2, and H5 moderately inhibited surfactant's surface activity. We also examined the cross-reactivity of these monoclonal antibodies with bovine (CLSE) and porcine (Curosurf) surfactants. By Western blot analysis, only H6 bound these heterologous surfactants. Other antibodies did so by ELISA. However, functional assays indicated that antibodies H7, H8 and H4 all greatly inhibited CLSE surface activity in vitro. Five antibodies (H1-H4 and H8) inhibited Curosurf function. Thus, human surfactant species, especially low molecular weight species, are highly antigenic. Antibodies to alveolar surfactants may inhibit surfactant function in vitro. As indicated by Western blot and cross-inhibition data, human lower molecular weight surfactants share epitopes with proteins from therapeutically important porcine and bovine surfactants. The potential importance of these findings to treatment of neonatal respiratory distress syndrome with heterologous surfactants is discussed.

Amniotic Fluid↗
PubMed1991

Immunogenicity of surfactant. II. Porcine and bovine surfactants.

Protein-containing surfactants of human and animal origin are being used increasingly to treat neonatal and adult respiratory distress syndromes. This trend led us to examine the antigenicity of two important preparations of animal surfactant, cow lung surfactant extract (CLSE) and a porcine surfactant preparation, Curosurf. We describe here 15 monoclonal antibodies against Curosurf and four against CLSE. Antibodies were studied by Western blot analysis to determine their ability to recognize protein components of their respective surfactant preparations. They were also tested for their ability to inactivate surfactant in vitro, assayed using the pulsating bubble surfactometer. Several antibodies directed against CLSE or Curosurf functionally inactivate the surfactant to which they were raised. We determined the degree of immunologic cross-reactivity between antibodies directed to CLSE and Curosurf against the other surfactant and also against human surfactant, both by Western blot and by examining functional inactivation in vitro. Antibodies to these animal surfactants that are commonly used therapeutically may inactivate the specific animal surfactant to which they were raised, as well as human and other surfactants. Generally, when antibodies inactivate surfactant from more than one animal species, they inactivate heterologous surfactants comparably to the extent to which they inactivate the surfactant to which they are directed. Immune complexes between anti-surfactant antibodies and surfactant have been described in the course of neonatal respiratory distress syndrome. The potential pathophysiological importance of anti-surfactant antibodies may therefore lie in their ability to inactivate administered surfactant, other similar surfactants and endogenous surfactant. In so doing, these antibodies may potentiate surfactant deficiency or pulmonary injury initiated by other stimuli.

Animals↗
PubMed1991

Effects of antisurfactant antibodies on the course of mild respiratory distress syndrome.

The role of surfactant-associated proteins in surfactant function was studied by selectively blocking these proteins with monoclonal antibodies. Four monoclonal antibodies, M1, M2, M3, and M4 were identified and their reactivities examined by Western blot analysis. M1, M2, and M4 bind, respectively, 8-, 10- and both 10- and 34-kD proteins. M3 antibody did not recognize a protein as assayed by this technique. These antibodies were then administered intratracheally to adult rats that had been partially depleted of lung surfactant by broncholavage. None of these antibodies had any deleterious effect on pulmonary function. On the other hand, M4 antibody significantly improved gas exchange. Possible mechanisms by which antibody may effect such improvement are discussed.

Animals↗
PubMed1991

In vivo and in vitro inactivation of bovine surfactant by an anti-surfactant monoclonal antibody.

In this study the importance of a low-weight surfactant protein (11 kDa) is demonstrated by selectively blocking this protein with a monoclonal antibody. In adult rats respiratory failure was induced by repeated bronchoalveolar lavage to remove all pulmonary surfactant. It was shown that surfactant mixed with the antibody was not capable of restoring lung function when compared with surfactant alone or surfactant mixed with control serum. Using the pulsating bubble surfactometer, it could be demonstrated that surfactant mixed with this antibody had a significant higher minimum surface tension when compared with surfactant alone, or surfactant mixed with an unrelated mouse immunoglobulin G (IgG). The inhibition of surfactant function by the monoclonal antibody suggests the importance of the 11 kDa protein for normal surfactant function.

Animals↗
PubMed1991

Identification of a cell membrane protein that binds alveolar surfactant.

Alveolar surfactants are complex mixtures of proteins and phospholipids produced by type II alveolar cells and responsible for lowering pulmonary surface tension. The process by which surfactant is produced and exported and by which its production by pulmonary cells is regulated are not well understood. This study was designed to identify a cellular receptor for surfactant constituents. To do so, monoclonal anti-idiotypic antibodies directed against antibodies to porcine and rabbit surfactant proteins were prepared. These monoclonal anti-idiotypic antibodies bind both alveolar lining and bronchial epithelial cells in rabbit, porcine, and human lungs. Macrophages and other nonepithelial cells do not react with these antibodies. Western blot analysis indicates that both A2R and A2C recognize the same proteins in both pig and rabbit lungs: a 30-kd protein and additional proteins at 52 and 60 kd. Preincubating lung wash cells with A2C or A2R prevents binding of porcine or rabbit surfactant preparations, respectively, by these cells. Preincubating frozen sections of lung tissue with surfactant inhibits binding of A2R and A2C to the lung. Antibody directed to a cell membrane protein that recognizes alveolar surfactant may be useful in elucidating the structure and function of this receptor and in understanding the cellular physiology and pathophysiology of the surfactant system.

Animals↗
PubMed1990

Molecular analysis of immunosuppression induced by virus replication in lymphocytes.

The relationship between immunosuppression and oncogenesis can be determined by studying the molecular interactions between tumor-inducing viruses and lymphocytes. We approached this study by using a unique system of two genetically related Leporipoxviruses, malignant fibroma virus (MV), and Shope fibroma virus (SFV). MV induces a syndrome of a highly lethal, disseminated myxosarcoma, severe immune suppression, and replicates in lymphocytes both in vivo and in vitro. In contrast, SFV causes a benign fibromyxosarcoma without immune dysfunction and cannot replicate in lymphocytes. Earlier studies demonstrated that transfer of a 10.8-kb Bam HI piece of MV (fragment "C") to SFV resulted in the ability of SFV to replicate in lymphocytes and suppress immune function. These results suggested that lymphocytotropic replication and immune suppression was located on the left side of fragment C. We extended these studies by generating families of recombinants between MV and SFV by using subfragments of fragment C. The resulting recombinant viruses were analyzed for their ability to replicate in lymphocytes, suppress immune function, and produce tumors. Those recombinants expressing MV-like characteristics were mapped by endonuclease digestion. This study demonstrates that recombinants containing a 3.6-kb Nde I subfragment, as well as those containing an overlapping 1.9-kb Hinc II subfragment, were capable of replicating in lymphocytes, suppressing immune functions, and inducing disseminated tumors in rabbits. Our study has therefore identified a portion of MV DNA sufficient to transfer the unique pathogenicity of MV to SFV, and suggests that control of immune suppression and tumor dissemination may not necessarily be mediated by the same viral genes.

Animals↗
PubMed1990

Natural cell-mediated immunity in the rabbit.

Susceptibility and resistance to tumors represent the interplay of many factors. One factor felt to govern the development of tumors is natural killer and natural cytotoxic cellular activity. The constitutional resistance of rabbits to spontaneous tumor development raises questions regarding the activity of natural cell-mediated immunity in this species. We therefore examined the ability of rabbit spleen, lymph node, and peripheral blood lymphocytes to mediate natural killer cell (NK) and natural cytotoxic cell (NC) activity in vitro. Using classical approaches to the study of NK and NC activity, we found no evidence of these activities in leporine spleen, lymph node, and peripheral blood lymphocytes. Preincubation of these cells with IL-2 did not induce such activity. Antibody-dependent cell-mediated cytotoxic reactivity (ADCC), which is believed to be mediated by NK cells, was also undetectable in rabbit lymphocytes. As controls, lymphocytes from other species were capable of mediating NK, NC, and ADCC functions normally in these experiments. Finally, we were unable to identify a population of large granular lymphocytes, the cells believed to mediate NK activity in other animals. Therefore, we could not demonstrate in the rabbit either natural cell-mediated immunity or the population of cells usually associated with natural cell-mediated immunity. If such activity exists in rabbits, it is different from that seen in other animals. More likely, the basis for the natural resistance of rabbits to tumor development must be sought elsewhere.

Animals↗
PubMed1990

Determinants of the ability of malignant fibroma virus to induce immune dysfunction and tumor dissemination in vivo.

The relationship of virus-induced immunological dysfunction and tumor dissemination was studied using two related tumor-causing leporipoxviruses: malignant fibroma virus (MV) and Shope fibroma virus (SFV). Recombinant viruses, produced by transferring MV's 10.7 kb BamHI C fragment to SFV, replicate in lymphocytes and suppress lymphocyte function in vitro. Those recombinants that replicate in lymphocytes and suppress lymphocyte function in vitro share about 3.5 kb from MV's C fragment. Some recombinants mimic MV in producing immune suppression and disseminated virus infection in vivo. Other recombinants, even some that are highly immunosuppressive in vitro (e.g. R71), only variably induce immune suppression in vivo, and do not cause disseminated disease. A segment of DNA from MV that transfers to Shope fibroma virus almost all of MV's virulence in vivo was identified.

Animals↗