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Biomedical subjects

J M Crawford

Publications and source records attributed to J M Crawford.

At least 19 recordsLinked to original sources
PubMed2005

Anaesthetists should wear gloves--touch sensitivity is improved with a new type of thin glove.

BACKGROUND: Sterile gloves should be worn for all invasive procedures. However, loss of touch sensitivity when wearing gloves leads to poor compliance among anaesthetists in the UK. Our aim was to investigate whether new surgical glove types, such as extra-thin or latex-free gloves, offer any advantage in touch sensitivity. METHODS: The skin-pressure sensibility threshold of the gloved pulp of the dominant index finger was established in 24 anaesthetists using nine self-made modified von Frey hairs. The range of forces generated by nylon monofilaments was evenly distributed between 0.5 and 17 mN. The gloves tested were latex Biogel standard, latex-free Biogel Neotech, and extra-thin latex Ansell microthin surgical gloves. The different glove types were assessed in a counterbalancing algorhithm, the individual was blinded, and the filaments were applied in random order. RESULTS: The median threshold force to touch was significantly lower for the extra-thin surgical gloves in comparison to the standard latex (P = 0.014) and latex-free gloves (P = 0.001). There was no significant difference between the standard latex and latex-free gloves (P = 0.166). CONCLUSION: We have demonstrated that wearing extra-thin surgical gloves offers increased touch sensitivity. This may improve dexterity when carrying out delicate invasive procedures. Latex-free surgical gloves performed similar to standard latex gloves. Our findings should encourage more widespread glove use among anaesthetists.

Anesthesia↗
PubMed2004

Immunity to adeno-associated virus serotype 2 delivered transgenes imparted by genetic predisposition to autoimmunity.

Adeno-associated virus (AAV) is widely considered a promising vector for therapeutic gene delivery. This promise is based on previous studies assessing AAVs safety and toxicity, ability to infect nondividing cells, elicit a limited immune response and provide long-term gene expression. However, we now find that earlier studies underappreciated the degree of AAV immunogenicity as well as the extent to which genetic background, through regulation of immune responsiveness, influences the duration of gene expression and thereby the effectiveness of AAV-mediated gene therapy. We evaluated antibody responses in 12 mouse strains to AAV serotype 2 (AAV2) and AAV2-expressed transgene products including green fluorescent protein (GFP), human alpha1-antitrypsin and murine interleukin-10. As expected, all immunocompetent mice administered AAV2 developed serologic evidence of immune responsiveness to the virus. However, a previously unidentified serologic prozone effect was observed suggesting that the concentrations of anti-AAV2 antibodies may have historically been subject to marked underestimation. Furthermore, strains with genetic predisposition to autoimmunity (eg, NOD, NZW, MRL-lpr) specifically imparted a functionally deleterious immune response to AAV-delivered transgene products. These findings suggest that more thorough studies of anti-AAV immunity should be performed, and that genetic predisposition to autoimmunity should be considered when assessing AAV efficacy and safety in humans.

Animals↗
PubMed2004

Recombinant adeno-associated virus-mediated alpha-1 antitrypsin gene therapy prevents type I diabetes in NOD mice.

Type I diabetes results from an autoimmune destruction of the insulin-producing pancreatic beta cells. Although the exact immunologic processes underlying this disease are unclear, increasing evidence suggests that immunosuppressive, immunoregulatory and anti-inflammatory agents can interrupt the progression of the disease. Alpha 1 antitrypsin (AAT) is a multifunctional serine proteinase inhibitor (serpin) that also displays a wide range of anti-inflammatory properties. To test the ability of AAT to modulate the development of type I diabetes, we performed a series of investigations involving recombinant adeno-associated virus vector (rAAV)-mediated gene delivery of human alpha-1 antitrypsin (hAAT) to nonobese diabetic (NOD) mice. Recombinant AAV-expressing hAAT (rAAV2-CB-AT) was administered intramuscularly to 4-week-old female NOD mice (1 x 10(10) i.u./mouse). A single injection of this vector reduced the intensity of insulitis, the levels of insulin autoantibodies, and the frequency of overt type I diabetes (30% (3/10) at 32 weeks of age versus 70% (7/10) in controls). Transgene expression at the injection sites was confirmed by immunostaining. Interestingly, antibodies against hAAT were present in a majority of the vector-injected mice and circulating hAAT was undetectable when assessed 10 weeks postinjection. This study suggests a potential therapeutic role for AAT in preventing type I diabetes as well as the ability of AAV gene therapy-based approaches to ameliorate disease effectively.

Animals↗
PubMed2004

The influence of esmolol on the dose of propofol required for induction of anaesthesia.

Cardiac output may be an important determinant of the induction dose of intravenous anaesthetic. Esmolol is known to reduce cardiac output, and we examined its effect on the propofol dose required for induction of anaesthesia. The size of the effect seen with esmolol was compared with midazolam co-induction. Sixty patients were randomly allocated to placebo (saline), esmolol (1mg x kg(-1) bolus, followed by an infusion at 250 microg x kg(-1)min(-1)) or midazolam (0.04 mg x kg(-1)) groups. Induction of anaesthesia commenced 3 min following the administration of the study drug, using a Diprifusor set to achieve plasma propofol concentrations of 10 microg x ml(-1) at 5 min. The primary end point used was the propofol dose per kg at loss of response to command. The mean (SD) propofol dose for each group was 2.38 (0.48) mg x kg(-1) for placebo, 1.79 (0.36) mg x kg(-1) for esmolol and 1.34 (0.35) mg x kg(-1) for midazolam (all means significantly different; p < 0.0005). We found that predosing with esmolol reduces the propofol requirements for induction of anaesthesia by 25%.

Adjuvants, Anesthesia↗
PubMed2004

Interobserver reliability between a nurse and anaesthetist of tests used for predicting difficult tracheal intubation.

We examined the interobserver reliability, between a nurse and anaesthetist, of five tests used to predict difficult tracheal intubation: mouth opening; thyromental distance; head and neck movement; mandibular luxation; and assessment of oropharyngeal view. For each test, an anaesthetic nurse and a specialist registrar anaesthetist were trained to use a standard method of examination. Most of the tests had either good or very good reliability. Assessment of mouth opening demonstrated only moderate reliability and assessment of oropharyngeal view demonstrated poor reliability. The interobserver reliability estimates between a nurse and an anaesthetist are similar to those previously demonstrated between two anaesthetists.

Head Movements↗
PubMed2003

Pre-operative screening: criteria for referring to anaesthetists.

Nurses assess patients pre-operatively using screening questionnaires and locally-developed protocols. Our objectives were to determine which questions might identify patients who should be seen by an anaesthetist before the day of surgery. A review of the literature and a preliminary questionnaire to establish questions to be tested was followed by a modified, two-round Delphi questionnaire to determine the level of agreement by anaesthetists. There was agreement for referring patients who gave a positive response to questions that query: restricted exercise tolerance; previous anaesthetic problems; family history of anaesthetic problem; pathology affecting neck movement; angina; arrhythmia; heart failure; asthma; epilepsy; insulin-dependent diabetes mellitus; liver disease and unspecified kidney disease. There was equivocal agreement on questions that report a myocardial infarction over one year ago, cerebrovascular accident, non insulin-dependent diabetes mellitus and thyroid disease. Nurses should use these criteria during pre-operative assessment to decide the timing of evaluation by an anaesthetist.

Anesthesiology↗
PubMed2003

Prevalence of ROPS--equipped tractors and farm/farmer characteristics.

Data were collected on tractor safety status and the prevalence of ROPS as part of the Ohio Farm Family Health and Hazard Surveillance Program (FFHHSP) "walk-around" hazard assessments of cash grain farms in central Ohio. Principal operator (PO) characteristics thought a priori to have a possible association with ROPS utilization included age, education, percent time spent farming, total years in agriculture, and previous training in occupational injury prevention. Farm characteristics of interest were farm size, acres of cropland in use, sales value during the past 12 months, and current number of farm workers. Descriptive and logistic regression analyses were conducted to evaluate potential associations between these factors and ROPS utilization patterns. Of 1,044 tractors encountered on 306 farms, 359 (34.4%) were equipped with ROPS, 29 (9.5%) had nothing but ROPS-equipped tractors, and 117 (38.2%) did not have any ROPS-equipped tractors. The absence/presence of ROPS-equipped tractors in our sample was found to vary by several factors, and over a wide range. Results of invariable logistic regression analyses (excluding the relatively small number of tractors manufactured after 1985) indicated that younger POs with at least a high school education who spent more than half their time working in agriculture, employed three or more workers, had 500 or more acres of total farm land and cropland in use, and had a 12-month sales value exceeding $250,000 were more likely to have at least one ROPS-equipped tractor on their farms. However, time spent farming, number of workers, and the 12-month sales value were not strongly predictive in multivariable models. Although more than 20% of the POs in our sample reported having had training in occupational injury prevention, this factor had essentially no value in predicting the absence/presence of ROPS-equipped tractors on their farms. The results of this study illustrate the need for interventions to encourage ROPS retrofitting that target POs with less education and who work on smaller and less productive farms. However, the low overall ROPS utilization rates suggest that all farmers could benefit from intervention efforts.

Accidents, Occupational↗
PubMed2001

Adeno-associated virus vector-mediated IL-10 gene delivery prevents type 1 diabetes in NOD mice.

The development of spontaneous autoimmune diabetes in nonobese diabetic (NOD) mice provides for their use as a model of human type 1 diabetes. To test the feasibility of muscle-directed gene therapy to prevent type 1 diabetes, we developed recombinant adeno-associated virus (rAAV) vectors containing murine cDNAs for immunomodulatory cytokines IL-4 or IL-10. Skeletal muscle transduction of female NOD mice with IL-10, but not IL-4, completely abrogated diabetes. rAAV-IL-10 transduction attenuated the production of insulin autoantibodies, quantitatively reduced pancreatic insulitis, maintained islet insulin content, and altered splenocyte cytokine responses to mitogenic stimulation. The beneficial effects were host specific, as adoptive transfer of splenocytes from rAAV IL-10-treated animals rapidly imparted diabetes in naive hosts, and the cells contained no protective immunomodulatory capacity, as defined through adoptive cotransfer analyses. These results indicate the utility for rAAV, a vector with advantages for therapeutic gene delivery, to transfer immunoregulatory cytokines capable of preventing type 1 diabetes. In addition, these studies provide foundational support for the concept of using immunoregulatory agents delivered by rAAV to modulate a variety of disorders associated with deleterious immune responses, including allergic reactions, transplantation rejection, immunodeficiencies, and autoimmune disorders.

Adjuvants, Immunologic↗
PubMed2001

An adenoviral system for tetracycline-regulated TGF-beta expression mediates a reversible cell cycle arrest.

The ability to regulate the proliferation of normal cells in a reversible manner would be a useful adjunct to some clinical therapies. including many types of cancer chemotherapy and surgery for periodontal regeneration. While the application of recombinant growth factors and cytokines to target cells is a logical approach to regulate cell proliferation, the high turnover rates of these peptide factors often make this approach impractical. Recombinant adenoviral vectors can be used to direct the expression of transgene products such as growth factors in many cell types in vitro and in vivo. We have adapted the tetracycline-regulated expression system to allow regulated transforming growth factor (TGF)-beta1 expression using recombinant adenovirus. We demonstrate that infection with a recombinant TGF-beta-encoding virus system in primary human oral keratinocytes and in a lung epithelial cell line is sufficient to allow a cell cycle arrest that is reversible upon tetracycline addition. This inhibition is efficient even after the infection of a minority of cells in a population. These results highlight the possibility of using low level infection with recombinant adenovirus to cause short-term blocks on cell proliferation.

Adenoviridae↗
PubMed2001

Stable therapeutic serum levels of human alpha-1 antitrypsin (AAT) after portal vein injection of recombinant adeno-associated virus (rAAV) vectors.

Previous work from our group showed that recombinant adeno-associated virus (rAAV) vectors mediated long-term secretion of therapeutic serum levels of human alpha-1 antitrypsin (hAAT) after a single injection in murine muscle. We hypothesized that hepatocyte transduction could be even more efficient, since these cells represent the natural site of AAT production and secretion. To test this hypothesis, rAAV vectors containing the hAAT cDNA driven by either the human elongation factor 1 alpha promoter, the human cytomegalovirus immediate-early promoter (CMV), or the CMV-chicken beta actin hybrid (CB) promoter were injected into the portal or tail veins of adult C57Bl/6 mice. Potentially therapeutic serum levels of hAAT (600 microg/ml) were achieved after portal vein injection of doses of 4 x 10(9) infectious units (IU), a 10-fold lower dose than that required for similar levels of expression via the i.m. route. Serum levels greater than 1 mg/ml were achieved at doses of 3 x 10(10) IU. Southern blotting of liver DNA revealed the presence of circular episomal vector genomes. Immunostaining showed that transgene expression was scattered throughout the liver parenchyma. Similar results were obtained with a rAAV-CB-green fluorescent protein (GFP) vector. There was no evidence of hepatic toxicity. These data indicate that liver-directed rAAV-based gene therapy is effective in the murine model, and hence might be feasible for treatment of human AAT deficiency.

Animals↗
PubMed2001

Observed incidence of tumorigenesis in long-term rodent studies of rAAV vectors.

Gene therapy using recombinant adeno-associated virus vectors (rAAV) is generally considered safe. During the course of a study designed to determine the long-term efficacy of rAAV-mediated gene therapy initiated in newborn mice with the lysosomal storage disease, mucopolysaccharidosis type VII (MPSVII), a significant incidence of hepatocellular carcinomas and angiosarcomas was discovered. A hepatocellular carcinoma was first detected in a 35-week-old mouse and by 72 weeks of age, three out of five rAAV-treated MPSVII mice had similar lesions. These types of tumors had not been seen previously in long-term studies of MPSVII mice using recombinant enzyme or bone marrow transplantation. In an attempt to ascertain whether mouse strain or GUSB expression confers susceptibility to tumor formation, we histopathologically examined untreated normal mice of the same strain, untreated MPSVII mice, and normal mice overexpressing human GUSB for the presence of tumors and increased hepatocyte replication. The results of these studies do not indicate that MPSVII mice or mice overexpressing human GUSB are susceptible to tumor formation; however, the number of animals examined is too small to draw definitive conclusions. Results from quantitative PCR performed on the tumor samples suggest that the tumors are probably not caused by an insertional mutagenesis event followed by the clonal expansion of a transformed cell. In a separate study, a relatively large group of mice injected with varying doses and types of rAAV vectors had no evidence of hepatic or vascular tumors. Although the mechanism of tumor formation is currently unknown, the tumorigenic potential of rAAV vectors must be rigorously determined in long-term in vivo studies.

Animals↗
PubMed2001

Role of myosin-II phosphorylation in V12Cdc42-mediated disruption of Drosophila cellularization.

Microinjection of constitutively active Cdc42 (V12Cdc42) disrupts the actomyosin cytoskeleton during cellularization (Crawford et al., Dev. Biol., 204, 151-164 (1998)). The p21-activated kinase (PAK) family of Ser/Thr kinases are effectors of GTP-bound forms of the small GTPases, Cdc42 and Rac. Drosophila PAK, which colocalizes with actin and myosin-II during cellularization, concentrates at sites of V12Cdc42-induced actomyosin disruption. In vitro biochemical analyses demonstrate that PAK phosphorylates the regulatory light chain (RLC) of Drosophila nonmuscle myosin-II on Ser21, a site known to activate myosin-II function. Although activated PAK does not disrupt the actomyosin cytoskeleton, it induces increased levels of Ser21 phosphorylated RLC. These findings suggest that increased levels of RLC phosphorylation do not contribute to disruption of the actomyosin hexagonal array.

Actomyosin↗
PubMed2001

Enhancement of survival by LPA via Erk1/Erk2 and PI 3-kinase/Akt pathways in a murine hepatocyte cell line.

First published September 5, 2001; 10.1152/ajpcell.00077.2001.-Protective mechanisms for lysophosphatidic acid (LPA) against cell death caused by Clostridium difficile toxin, or tumor necrosis factor-alpha (TNF-alpha) plus D-galactosamine, were investigated in a murine hepatocyte cell line AML12 expressing Edg2 LPA receptor. In these models of hepatocellular injury, LPA prevented hepatocyte damage, suppressed apoptosis, and enhanced cell survival in a dose-dependent fashion. The protective effects of LPA were abolished by wortmannin and LY-294002, specific inhibitors of phosphatidylinositol 3-phosphate kinase (PI 3-kinase), and by PD-98059 and U-0126, inhibitors of MEK1/MEK2. In nontreated hepatocytes, LPA elicited a gradual and sustained increase in phosphorylation of Erk1/Erk2 over 180 min of stimulation and downstream phosphorylation of p90RSK and transcription factor Elk-1. In C. difficile toxin-treated cells, LPA-induced phosphorylation of Erk1/Erk2 was rapid but transient, while p90RSK and Elk-1 phosphorylation did not change significantly. LPA stimulated phosphorylation of Akt in a time-dependent manner in both intact and toxin-treated AML12 hepatocytes. Wortmannin and LY-294002 abolished phosphorylation of Akt, further supporting activation of PI 3-kinase/Akt as a signaling pathway, which mediates hepatocyte protection by LPA. Taken together, these results demonstrate that LPA prevents cell apoptosis induced by C. difficile toxin and TNF-alpha/D-galactosamine in the AML12 murine hepatocyte cell line. Cell protection by LPA involves activation of the mitogen-activated protein kinase Erk1/Erk2 cascade and PI 3-kinase-dependent phosphorylation of Akt.

Androstadienes↗
PubMed2001

LPS antagonism reduces graft-versus-host disease and preserves graft-versus-leukemia activity after experimental bone marrow transplantation.

Acute graft-versus-host disease (GVHD) and leukemic relapse remain the two major obstacles to successful outcomes after allogeneic bone marrow transplantation (BMT). Recent studies have demonstrated that the loss of gastrointestinal tract integrity, and specifically the translocation of LPS into the systemic circulation, is critical to the induction of cytokine dysregulation that contributes to GVHD. Using a mouse BMT model, we studied the effects of direct LPS antagonism on GVHD severity and graft-versus-leukemia (GVL) activity. Administration of B975, a synthetic lipid-A analogue from day 0 to day +6, reduced serum TNF-alpha levels, decreased intestinal histopathology, and resulted in significantly improved survival and a reduction in clinical GVHD, compared with control-treated animals. Importantly, B975 had no effect on donor T cell responses to host antigens in vivo or in vitro. When mice received lethal doses of P815 tumor cells at the time of BMT, administration of B975 did not impair GVL activity and resulted in significantly improved leukemia-free survival. These findings reveal a critical role for LPS in the early inflammatory events contributing to GVHD and suggest that a new class of pharmacologic agents, LPS antagonists, may help to prevent GVHD while preserving T cell responses to host antigens and GVL activity.

Animals↗
PubMed2001

Negotiated safety and other agreements between men in relationships: risk practice redefined.

This study examines patterns of agreement, knowledge and practice which can prevent or facilitate HIV transmission among men who are in regular ('primary') male-to-male relationships. Data are from a national volunteer phone-in survey of homosexually-active men in Australia. A sub-sample of 1070 men from a larger sample (n=3039) were found to have one or more regular partners for longer than 6 months. Self-reported HIV serostatus of survey participant and his regular partner, type of agreement regarding anal intercourse both within and outside the regular relationship, and engaging in unprotected anal intercourse with regular and with casual partners were examined. Risk practice was defined as unprotected anal intercourse with a regular partner of different or unknown serostatus and/or unprotected anal intercourse with a casual partner. Agreements were classified as: negotiated safety (28.8%); no unprotected anal intercourse (33.6%); unsafe (10.6%) and no agreement (17.0%). Risk practice was reported by 17.8% of the men. Type of agreement was found to be the strongest predictor of risk practice. Negotiated safety agreements were common, and were kept on the whole. Men with unsafe agreements, although only a small proportion of men in relationships, had high levels of risk practice.

Australia↗
PubMed2000

Reduced susceptibility of nonobese diabetic mice to TNF-alpha and D-galactosamine-mediated hepatocellular apoptosis and lethality.

Nonobese diabetic (NOD/LtJ or NOD) mice are resistant to doses of LPS and D-galactosamine that uniformly produce lethality in C57BL/6J (B6) mice (p < 0.01). Liver caspase-3-like activity, serum transaminase levels (both p < 0.05), and the numbers of apoptotic liver nuclei were also reduced in NOD compared with B6 mice treated with LPS (100 ng) and D-galactosamine (8 mg). NOD mice were also at least 100-fold more resistant to recombinant human TNF-alpha and D-galactosamine treatment than B6 mice (p < 0.001). Binding of recombinant human TNF-alpha to splenocytes from NOD mice was similar to that seen in B6 mice, suggesting that the defect in responsiveness was not due to an inability of recombinant human TNF-alpha to bind the NOD TNF type 1 (p55) receptor. Because the TNF type 1 (p55) receptor shares a common signaling pathway with Fas (CD95), NOD and B6 mice were treated with the Fas agonist antibody, Jo-2. Surprisingly, NOD mice were as sensitive as B6 mice to Fas-induced lethality and hepatic injury. In addition, primary hepatocytes isolated from NOD mice and cultured in vitro in the presence of D-galactosamine with or without TNF-alpha were found to be resistant to apoptosis and cytotoxicity when compared with B6 mice. In contrast, Jo-2 treatment produced similar increases in caspase-3 activity and cytotoxicity in primary hepatocytes from NOD and B6 mice. The resistance to LPS- and TNF-alpha-mediated lethality and hepatic injury in D-galactosamine-sensitized NOD mice is apparently due to a post-TNFR binding defect, and independent of signaling pathways shared with Fas.

Animals↗