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S Craig

Publications and source records attributed to S Craig.

At least 55 records · Page 3Linked to original sources
PubMed1993

Characterization of the quaternary structure and conformational properties of the human stem cell inhibitor protein LD78 in solution.

The human LD78 protein (sometimes referred to as human macrophage inflammatory protein-1 alpha) has been shown to protect multipotential hemopoietic stem cells from the effects of cytotoxic agents. Administration of the recombinant stem cell inhibitor molecule LD78 as an adjunct to chemotherapy has potential clinical benefit in reducing or preventing the neutropenia associated with this treatment. At physiological ionic strength, the 8-kDa LD78 molecule exists as soluble, heterogeneous, multimeric complexes of mass ranging from 100 to > 250 kDa. The hydrodynamic and structural properties of LD78 have been determined in various buffer solutions using analytical ultracentrifugation, circular dichroism, and fluorescence spectroscopy. The results demonstrate that defined, homogeneous monomer and tetramer forms of LD78 can be prepared which display distinct conformational properties. The combined use of hydrodynamic and spectroscopic analysis provides an insight into the pathway and molecular mechanics of LD78 self-association.

Acetates↗
PubMed1993

Project towards no tobacco use: 1-year behavior outcomes.

OBJECTIVES: We present 1-year follow-up data from a school-based tobacco use prevention project designed to test the effectiveness of three main components of social influence programs. The components teach refusal skills, awareness of social misperceptions about tobacco use, and misconceptions about physical consequences. METHODS: Four different curricula were developed and tested in a randomized experiment involving 48 junior high schools. The outcome variables examined were changes in initial and weekly cigarette and smokeless tobacco use 1 year after the intervention. RESULTS: Analyses indicated that each of the component programs were effective in decreasing both the initial and the weekly use of cigarettes except for the curriculum in which refusal skills were taught. Also, each curriculum was effective in decreasing the initial use of smokeless tobacco except for the one aimed at correcting social misperceptions. Only the combined curriculum showed an effect on the weekly use of smokeless tobacco. CONCLUSIONS: The combined intervention was the most effective overall in reducing the initial and weekly use of cigarettes and smokeless tobacco. This suggests that different reasons for use exist and need to be counteracted simultaneously. However, since single programs were also effective in reducing all but weekly smokeless tobacco use, any of these components may be worthwhile prevention tools.

Adolescent↗
PubMed1993

Which lesson components mediate refusal assertion skill improvement in school-based adolescent tobacco use prevention?

Any of three components of current school-based refusal assertion training might mediate improvement of seventh grade students' ability to refuse tobacco use offers: 1) teaching students knowledge of ways to say "no," 2) engaging students in the practice of refusal assertion, or 3) motivating students to perform refusal assertion in a socially skilled way. A 3-condition true field experimental "component study" of the differential effects of these three components yielded improvement in role-played behavioral skill to refuse tobacco offers that was evident in both the Knowledge and Practice conditions but not in the Motivation condition. In these same two conditions, skills training led to a significant decrease in students' intention to use smokeless tobacco in the future but not cigarettes. A focus on engaging students in Knowledge and Practice components of refusal assertion training appears warranted.

Adolescent↗
PubMed1992

Expression and genomic structure of the genes encoding FdI, the major allergen from the domestic cat.

The genes encoding chain 1 (Ch1) and chain 2 (Ch2) of the major allergen of the domestic cat, Felis domesticus I, have been analyzed by genomic cloning and by polymerase chain reaction (PCR). Ch1 has two potential leader sequences, designated A and B. Analysis of a genomic clone encoding Ch1 demonstrated that one structural gene contains sequences corresponding to both leaders, which utilize different Met start codons. PCR analysis showed that genes encoding Ch1 and Ch2 are co-expressed in both the salivary glands and the skin, and that leader sequence A of Ch1 is utilized preferentially in both tissues. Ch2 was shown to have two dominant forms that are differentially expressed in the aforementioned tissues. The long form (Ch2L), composed of 92 amino acids (aa), is preferentially expressed in the salivary glands, while the short form (Ch2S), composed of 90 aa, is preferentially expressed in the skin. There is minor sequence polymorphism in both forms of Ch2. A genomic clone for Ch2 only contained sequences for Ch2S, suggesting that Ch2L is encoded by an exon not contained within this genomic clone.

Allergens↗
PubMed1992

Purification and analysis of proteinase-resistant mutants of recombinant platelet-derived growth factor-BB exhibiting improved biological activity.

Recombinant platelet-derived growth factor (PDGF)-BB was expressed and secreted from yeast in order to study the structure-function relationships of this mitogen. A simple purification scheme has been developed which yields greater than 95% pure PDGF-BB. Analysis of this recombinant PDGF-BB shows partial proteolysis after arginine-32. Substitution of this arginine residue, or arginine-28 [a potential KEX2 (lysine-arginine endopeptidase) cleavage site], prevents or reduces cleavage of PDGF-BB respectively. These mutations result in a 5-fold increase in expression levels of PDGF-BB, and the resulting mutant proteins show higher activity in a number of biological assays than the cleaved wildtype PDGF-BB. These data are in accord with previous work by Giese, LaRochelle, May-Siroff, Robbins & Aaronson [(1990) Mol. Cell Biol. 10, 5496-5501] suggesting that the region isoleucine-25-phenylalanine-37 is involved in PDGF-receptor binding.

3T3 Cells↗
PubMed1992

Characterization of the structure and conformation of platelet-derived growth factor-BB (PDGF-BB) and proteinase-resistant mutants of PDGF-BB expressed in Saccharomyces cerevisiae.

A detailed biophysical study of the secondary and tertiary structures of recombinant platelet-derived growth factor (PDGF)-BB produced in yeast has been carried out. The secondary structure of the molecule is composed of 54% beta-sheet with less than 5% ordered helix. The single tryptophan residue has been shown to be solvent-accessible; however, the ability of the side chain to rotate is severely restricted. The fluorescence emission is quenched at pH 7.0 and in the presence of high salt, but dequenched by titration to lower pH with a pK of 5.8. Two proteinase-resistant mutants of PDGF [( Ser28]- and [Pro32]-PDGF-BB) have also been characterized and shown to have secondary and tertiary structures indistinguishable from wild-type PDGF-BB. These are, therefore, suitable stable background molecules in which to carry out structure-activity-relationship studies on PDGF-BB.

Mutagenesis, Site-Directed↗
PubMed1992

Biological and structural properties of MIP-1 alpha expressed in yeast.

The murine macrophage inflammatory proteins-1 alpha (MIP-1 alpha) and MIP-1 beta are distinct but closely related cytokines. Partially purified mixtures of the two proteins affect neutrophil function and cause local inflammation and fever. The particular properties of MIP-1 alpha have not been well studied, although it has been identified as being identical to an inhibitor of haemopoietic stem cell growth. We have expressed MIP-1 alpha in yeast cells and purified it to sequence homogeneity. Structural analysis of this biologically active material by circular dichroism and fluorescence spectroscopy confirms that MIP-1 alpha has a very similar secondary and tertiary structure to platelet factor 4 and interleukin 8 with which it shares limited sequence homology. The in-vitro stem cell inhibitory properties have been confirmed using a range of murine progenitor cells including purified bone marrow progenitor cells (FACS-1), the FDCP-mix A4 cell line, and spleen colony forming unit (CFU-S) populations. Plateau levels of inhibition of stem cell growth were achieved using concentrations of 0.15 micrograms/ml MIP-1 alpha. We have also demonstrated that MIP-1 alpha is active in vivo: 5 micrograms of MIP-1 alpha per mouse given as a bolus injection, protects stem cells from subsequent in-vitro killing by tritiated thymidine. MIP-1 alpha was also shown to enhance the proliferation of more committed progenitor granulocyte macrophage-colony forming cells (GM-CFC) in response to granulocyte macrophage-colony stimulating factor (GM-CSF).

Animals↗
PubMed1992

Vicilin with carboxy-terminal KDEL is retained in the endoplasmic reticulum and accumulates to high levels in the leaves of transgenic plants.

Gene constructs were designed to test the effect of the endoplasmic reticulum (ER)-targeting signal, KDEL, on the level of accumulation of a foreign protein in transgenic plants. The gene for the pea seed protein vicilin was modified by the addition of a sequence coding for this tetrapeptide at its carboxyl terminus. The altered gene was placed under the control of a CaMV 35S promoter and its expression in the leaves of both tobacco and lucerne (alfalfa) was compared with that of an equivalent vicilin construct lacking the KDEL-coding sequence. The presence of the ER-targeting signal led to a greatly enhanced accumulation of the heterologous protein. In lucerne and tobacco leaves, the level of vicilin-KDEL protein was 20 and 100 times greater than that of the unmodified vicilin, respectively. These differences in expression level could not be explained by corresponding differences in the steady-state levels or the translatability of the mRNAs. However, when the stability of vicilin and vicilin-KDEL proteins was compared in their respective transgenic hosts, unmodified vicilin was found to be degraded with a half-life of 4.5 h while vicilin-KDEL was much more stable with a half-life of more than 48 h. Immunogold labelling of leaf tissues from transgenic lucerne and tobacco showed the presence of vicilin associated with large aggregates within the ER lumen of vicilin-KDEL plants. No such aggregates were detected in transgenic plants expressing wild-type vicilin. It is concluded that the carboxy-terminal KDEL caused the retention of the modified vicilin in the ER, and that this retention led to the increased stability and higher level of accumulation of vicilin-KDEL in leaves of transgenic plants.

Amino Acid Sequence↗
PubMed1992

Ultraviolet A irradiation stimulates collagenase production in cultured human fibroblasts.

This study was designed to investigate the biochemical mechanisms responsible for the connective tissue changes seen in actinically damaged skin, which is characterized histologically by diminution and ultrastructural alterations of collagen fibrils and deposition of elastotic material in the papillary dermis. We hypothesized that ultraviolet light could stimulate synthesis of interstitial collagenase in the skin, resulting in collagen degradation. Monolayer cultures of human fibroblasts or keratinocytes were irradiated with ultraviolet A (UVA) or ultraviolet B (UVB) radiation and interstitial collagenase or its inhibitor, TIMP (tissue inhibitor of metalloproteinases) assessed in the conditioned medium with Western immunoblots 24 h after irradiation. Northern blot analysis of the irradiated fibroblasts with a cDNA probe representing collagenase was also performed. Cell viability was greater than 90% with all doses of UV radiation studied. A dose-related increase in immunoreactive collagenase was detected in the medium of fibroblasts irradiated with 0-10 J/cm2 of UVA radiation as well as a parallel increase in the collagenase mRNA in the irradiated cells. UVA radiation stimulated collagenase synthesis in both neonatal and adult fibroblasts. TIMP production in UVA-irradiated fibroblasts increased to a lesser degree than did collagenase and its increase did not parallel the increase in collagenase. UVB (0-100 mJ/cm2) did not stimulate collagenase production by fibroblasts. In contrast to the stimulation of collagenase production by fibroblasts, a slight decrease in immunoreactive collagenase was seen in UVA-irradiated keratinocytes. These data suggest that direct stimulation of collagenase synthesis by human skin fibroblasts by UVA radiation may contribute to the connective tissue damage induced by ultraviolet radiation leading to photoaging.

Cells, Cultured↗
PubMed1992

ORSOS goes to theatre!

The presence of computers in the operating theatre is on the increase. Sheena Craig, theatre manager at the Western General Hospital, Edinburgh, describes how the Operating Room Scheduling and Office System (ORSOS) was installed in her theatre suite and asks, 'Was it worth the effort'?!

Attitude of Health Personnel↗
PubMed1991

Two PDGF-B chain residues, arginine 27 and isoleucine 30, mediate receptor binding and activation.

PDGF may be involved in the pathogenesis of a variety of disorders including atherosclerosis and certain types of cancer. There is currently little understanding of the molecular structure of PDGF and of the critical amino acid residues involved in receptor binding and cell activation. Two such PDGF-B chain residues, arginine 27 and isoleucine 30, have been identified by a site-directed mutagenesis programme. Substitutions in these positions can lead to PDGF mutants defective in both receptor affinity and cell activation as judged by displacement of [125I]PDGF-BB, mitogenic assay and inositol lipid turnover. Circular dichroism and fluorescence spectroscopy show that such mutations do not disrupt the structure of PDGF.

3T3 Cells↗
PubMed1991

Restriction fragment length polymorphisms of the apolipoprotein A-I, C-III, A-IV gene locus. Relationships with lipids, apolipoproteins, and premature coronary artery disease.

Data from various laboratories have indicated associations of various alleles determined by RFLPs within or adjacent to several apolipoprotein genes with abnormalities in plasma lipids and/or premature coronary artery disease (CAD). In order to assess such relationships we have examined allele frequencies of 8 different RFLPs within or adjacent to the apo A-I, C-III and A-IV gene complex on the long arm of chromosome 11 (MspI, 5' to the apo A-I gene; MspI, within the apo A-I gene; PstI, 3' to the apo A-I gene; SstI, 3' to the apo C-III gene; PvuII, within the apo C-III gene; PvuII, 5' to the apo C-III gene; XbaI, within the apo A-IV gene; and XbaI, 3' to the apo A-IV gene) in 202 patients with CAD (50% narrowing of one or more coronary arteries) prior to age 60 and 145 normal controls. None of the allele frequencies of these RFLPs were significantly different in cases as compared to controls. With regard to associations with plasma lipids and apolipoprotein levels, the rare allele determined by the absence of the PstI site was associated with elevated triglyceride levels (P less than 0.05) in cases, but not in controls. In contrast, the rate MspI allele 5' to the apo A-I gene was associated with elevated triglyceride levels (P less than 0.05) in controls but not in cases. In both cases and controls, subjects with the uncommon SstI allele had triglyceride levels that were 9 and 38% higher than in those without this allele. These differences were significant (P less than 0.05) only in controls. Our data indicate that the rare allele determined by the SstI site within this gene complex deserves further study in order to understand its association with elevated triglycerides in Caucasian populations. However, at the present time all these DNA markers lack sufficient specificity to be clinically useful for CAD risk assessment.

Age Factors↗
PubMed1990

Purification and secondary structure determination of simian immunodeficiency virus p27.

We have developed a novel method for the expression and purification of p27, the major core protein of simian immunodeficiency virus. Circular dichroism measurements of purified p27 were used to determine the relative amounts of alpha-helix, beta-sheet and unordered secondary structural elements. These empirically determined values appear to be inconsistent with previously published theoretical models based on homology comparisons.

Circular Dichroism↗
PubMed1990

Peer-group association and adolescent tobacco use.

Mosbach & Leventhal (1988) examined the relation of cigarette smoking to peer-group identification in rural Wisconsin adolescents. They found that among dirts (problem-prone youth), regulars (average youth), hot-shots (good social or academic performers), and jocks (athletes), youth most likely to smoke were dirts and hot-shots. We performed a replication with a Southern California cohort and also for use of smokeless tobacco. We hypothesized that jocks would be the main users of smokeless tobacco. We identified the same groups and an additional one, skaters (skateboarders or surfers). As Mosbach & Leventhal found, cigarettes were used most by dirts. Contrary to their results, but consistent with other research, we found that hot-shots were least likely to smoke. Contrary to our prediction, we found that skaters and dirts were more likely to use smokeless tobacco than were jocks. Our data show that both tobacco forms are used by problem-prone youth.

Adolescent↗
PubMed1990

Nonlegume hemoglobin genes retain organ-specific expression in heterologous transgenic plants.

Hemoglobin genes from the nitrogen-fixing nonlegume Parasponia andersonii and the related non-nitrogen-fixing nonlegume Trema tomentosa have been isolated [Landsmann et al. (1986). Nature 324, 166-168; Bogusz et al. (1988). Nature 331, 178-180]. The promoters of these genes have been linked to a beta-glucuronidase reporter gene and introduced into both the nonlegume Nicotiana tabacum and the legume Lotus corniculatus. Both promoters directed root-specific expression in transgenic tobacco. When transgenic Lotus plants were nodulated by Rhizobium loti, both promoter constructs showed a high level of nodule-specific expression confined to the central bacteroid-containing portion of the nodule corresponding to the expression seen for the endogenous Lotus leghemoglobin gene. The T. tomentosa promoter was also expressed at a low level in the vascular tissue of the Lotus roots. The hemoglobin promoters from both nonlegumes, including the non-nodulating species, must contain conserved cis-acting DNA signals that are responsible for nodule-specific expression in legumes. We have identified sequence motifs postulated previously as the nodule-specific regulatory elements of the soybean leghemoglobin genes [Stougaard et al. (1987). EMBO J. 6, 3565-3569].

Base Sequence↗