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J M Wilson

Publications and source records attributed to J M Wilson.

At least 271 records · Page 15Linked to original sources
PubMed1995

Adenovirus-mediated gene transfer to human bronchial submucosal glands using xenografts.

The cystic fibrosis (CF) transmembrane conductance regulator has been localized to both submucosal glands and surface epithelium, suggesting that both glandular and surface epithelium may be important targets for gene therapy. To determine the distribution and efficiency of recombinant adenovirus-mediated gene transfer to human airway submucosal glands, an in vivo model was developed by heterotopically transplanting human bronchial segments from both normal and CF lung tissue into severe combined immunodeficient mice. A serotype 5 E1-deleted recombinant adenovirus containing a lacZ reporter gene driven by the cytomegalovirus promoter (H5.010CMVlacZ) was used to infect the xenografts. Transgene expression was correlated with three factors: 1) viral titer, 2) penetration of microspheres, and 3) dwell time of the viral instillate. At viral titers ranging from 10(8) to 10(11) plaque forming units/ml, expression of the lacZ gene was observed in a subpopulation of epithelial cells within approximately 40% of submucosal glands, with more efficient gene transfer to the ducts than the secretory tubules. Within individual glands, gene transfer varied from < 1% to approximately 20% of submucosal cells, including duct, mucous, and serous cells. Adenovirus-sized fluorescent microspheres were found to penetrate only some of the submucosal glands, suggesting that the gene transfer efficiency to submucosal tubules is due to limited viral particle penetration rather than tropism. Gene transfer to surface epithelial cells was inefficient. However, the percentage of transduced surface epithelial cells increased from < 1% to 5-10% as the dwell time of viral solution was increased from 5 min to 1 h, indicating that the time allowed for virus binding and entry is important for gene transfer efficiency.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcysteine↗
PubMed1995

Adenovirus-mediated gene transfer to human fetal lung ex vivo.

Gene therapy of the fetus or newborn infant is a potentially useful approach for prevention or treatment of specific lung diseases. To begin to address issues such as efficiency, duration, and cellular distribution of transgene expression, we studied transduction of human lung cells by recombinant, replication-deficient adenovirus containing the lacZ gene driven by the beta-actin promoter and cytomegalovirus enhancer (H5.010CBlacZ). Human fetal lungs of 20- to 24-wk gestation received approximately 10(11) viral particles by instillation into a major bronchus, and the tissue was cultured as explants in serum-free Waymouth's medium. beta-galactosidase staining (X-gal) was detected by 24 h in defined regions of treated tissue and localized to epithelial cells of airways and terminal saccules. beta-galactosidase activity in homogenate of treated tissue was maximal 3-5 days after exposure to virus, ranging from 0.2 to 1.5 A420.min-1.mg protein-1 in four experiments (control values were approximately 0.001). When virus was added directly to lung explants in culture, beta-galactosidase was expressed in most of the peripheral cells and rarely in interior cells, the level of activity was dose dependent between 10(8) and 10(11) viral particles/ml, and transgene expression was sustained for at least 28 days. Treatment of isolated cultured cells with virus resulted in equivalent staining of both epithelial cells and fibroblasts. We conclude that fetal lung cells are efficiently transduced by recombinant adenovirus, indicating the feasibility of gene therapy in the infant or fetus. Cultured fetal lung may be useful for testing gene constructs being considered for therapy.

Adenoviridae↗
PubMed1995

ICAM-1 expression on bronchial epithelium after recombinant adenovirus infection.

Early experience with recombinant adenoviruses for gene transfer to airway epithelium suggests that these vectors are associated with the development of inflammation. The mechanisms for this are unclear, but previous work has shown that respiratory viruses can cause increased expression of intercellular adhesion molecule-1 (ICAM-1) on airway epithelial cells. We therefore hypothesized that recombinant adenoviruses may induce ICAM-1 expression and thereby facilitate the development of airway inflammation. To address this, primary cultures of human bronchial epithelial cells were examined for ICAM-1 expression by flow cytometry after infection with a serotype 5, E1/E3-deleted recombinant adenovirus containing the Escherichia coli LacZ reporter gene driven by the cytomegalovirus promoter (Ad.CMVlacZ). Compared with control cells, ICAM-1 expression was unchanged after infection with Ad.CMVlacZ, but increased after infection with wild-type adenovirus. Treatment of Ad.CMVlacZ-infected cells with interferon-gamma (IFN) resulted in increased ICAM-1 expression, but to a lower level than that seen in cells treated with IFN alone, indicating that recombinant adenovirus infection blunted IFN-induced up-regulation of ICAM-1. Adhesion of human leukocytes to human bronchial epithelial cells was not increased after Ad.CMVlacZ infection, thereby excluding an ICAM-1-independent increase in leukocyte-epithelial adhesion. The results for ICAM-1 expression were confirmed in vivo, as immunostaining of human bronchial xenografts infected with Ad.CMVlacZ revealed basilar epithelial staining with ICAM-1, but no increased expression on cells expressing beta-galactosidase. This study demonstrates that unlike other respiratory viruses, recombinant E1/E3-deleted adenovirus does not cause increased ICAM-1 expression on human bronchial epithelium in vitro or in vivo nor increased leukocyte adhesion in vitro.

Adenoviridae↗
PubMed1995

Results of a national survey on anticoagulation for PTCA.

In order to assess the current practice patterns in the United States for anticoagulation during PTCA, a survey was sent to the ACC membership. A total of 377 surveys from physicians performing PTCA were tabulated; 5 (1.3%) respondents performed < 20 PTCAs/year, 128 (34.0%) performed 20-75 PTCAs/year, 141 (37.4%) performed 75-150 PTCAs/year, 98 (26.0%) performed > 150 PTCAs/year and 5 (1.3%) did not report their volume. Seventy-eight (20.7%) were at university hospitals 153 (40.6%) were at other teaching hospitals, 142 (37.7%) were at non-teaching hospitals and 4 (1.1%) did not report their institutional affiliation. A total of 76.8% of respondents routinely started with a 10,000 U bolus of heparin, while only 3.2% of respondents used only a weight-adjusted heparin bolus. Fifty-nine percent of respondents routinely used intra-procedure heparin infusions, usually 1000 U/hr. Anticoagulation monitoring was used by 92.6% of respondents during PTCA, almost always activated clotting times (ACTs). Of the 335 physicians who used ACTs to guide heparin therapy during PTCA, 59.1% used the Hemochron device, 16.7% used the HemoTec device, and 24.2% did not know which machine they used. Lower volume operators and operators at non-teaching hospitals were more likely not to know the type of ACT machine used. Post-procedure heparin infusions (usually titrated to an aPTT > 2 x control) were used by 70.3% of respondents. Lower volume operators were more likely to use post-procedure heparin infusions. Thus, heparin therapy for PTCA continues to be largely empiric, although the vast majority of cardiologists surveyed use ACT-guided heparin therapy for the procedure.(ABSTRACT TRUNCATED AT 250 WORDS)

Angioplasty, Balloon, Coronary↗
PubMed1995

A recombinant adenoviral vector expressing a soluble form of VCAM-1 inhibits VCAM-1/VLA-4 adhesion in transduced synoviocytes.

Intra-articular injection of recombinant adenovirus has been shown to be a feasible approach to the introduction of genetic reagents into synovial tissues in vivo. Rheumatoid arthritis (RA) is an autoimmune disorder characterized by the infiltration of lymphocytes and monocytes into inflamed synovium. It has been hypothesized that the recruitment of T lymphocytes/monocytes into sites of chronic inflammation is mediated by enhanced binding of very late antigen-4 (VLA-4) to vascular cell adhesion molecule-1 (VCAM-1) expressed on microvascular endothelial cells. Additional evidence suggests that VLA-4 binding continues to be important within the inflamed synovial membrane, where it appears to play a role in T cell retention and activation. A feasible therapeutic strategy for RA could be to utilize a soluble congener of the VCAM-1 molecule to block VLA-4 binding. In order to test this concept, a recombinant serotype Ad5 human adenovirus encoding a secreted form of VCAM-1 (Ad.CBsVCAM) was constructed. Human synoviocytes were readily infected in vitro with Ad.CBsVCAM, and sVCAM-1 expression and processing were analyzed by immunoprecipitation studies. Secretion of transgenic sVCAM was identified by ELISA of tissue culture supernatants, and biological activity was demonstrated with cell adhesion assays. In vivo, transgenic sVCAM-1 expression was determined by immunohistochemical analysis and in situ hybridization of synovial tissue, and secretion of transgenic sVCAM-1 was demonstrated by ELISA of tidal knee lavage fluid. The results showed that recombinant adenovirus can mediate the expression of a biologically active sVCAM-1 by synoviocytes in vivo and suggest that this strategy may be useful for inhibiting T lymphocyte retention and activation within rheumatoid synovium.

Adenoviridae↗
PubMed1995

Retrovirus-mediated expression of HUG Br1 in Crigler-Najjar syndrome type I human fibroblasts and correction of the genetic defect in Gunn rat hepatocytes.

Crigler-Najjar syndrome type I (CN-I) is a congenital hepatic metabolic deficiency in bilirubin UDP-glucuronosyltransferase activity which leads to profound jaundice and death from kernicterus. UGT1, the gene locus coding for multiple glucuronosyltransferase isoforms, has been well characterized and the cDNA for the most active form, HUG Br1, has been cloned. Recent advances in liver directed gene transfer suggest that this disease could be treated through gene therapy. As an initial step to correct the genetic defect in Crigler-Najjar type I, recombinant retroviruses were used to transduce an HUG Br1 gene into hepatocytes of a rat model of CN-I and CN-I fibroblasts. The retroviral vector gagCMVBA HUG Br1 was constructed and helper-free amphotrophic virus was isolated and used to transfer bilirubin UDP-glucuronosyltransferase activity to genetically deficient cells. The efficiency of transduction as measured by Southern blot analysis of integrated proviral sequences in DNA of recipient cells ranged from 5 to 100%. HUG Br1 gene expression was documented by blot hybridization analysis of total cellular RNA, immunotransblot analysis using a rabbit polyclonal antipeptide HUG Br1 antibody, and lysate enzymatic assay of bilirubin UDP-glucuronosyltransferase activity. HUG Br1 gene transfer was definitively demonstrated by four independent modalities following HUG Br1 retroviral transduction.

Amino Acid Sequence↗
PubMed1995

Revascularization therapy for coronary artery disease. Coronary artery bypass grafting versus percutaneous transluminal coronary angioplasty.

Coronary artery bypass surgery relieves the symptoms of myocardial ischemia and prolongs survival of patients with more severe coronary artery disease. Randomized trials of surgical therapy have consistently shown that the benefits of surgical revascularization are proportional to the amount of myocardium affected by, or at risk for, ischemic injury. This risk is inferred from angiographically delineated coronary anatomy, estimates of left ventricular function, and physiologic testing. The population that may see a survival benefit from surgical revascularization has probably been expanded beyond that reported in the VA, CASS, and ECSS trials, due to improved perioperative care, longer graft survival, and the use of internal mammary artery grafts. Percutaneous transluminal coronary angioplasty revascularizes myocardium by dilating a stenotic segment of coronary artery. While successful in relieving the symptoms of myocardial ischemia, PTCA is hindered by the occurrence of abrupt vessel closure and the frequent development of restenosis. Furthermore, firm proof of a survival benefit, outside of emergency therapy for acute myocardial infarction, is not yet available. However, because the risk of procedure-related death or serious complication is lower than that seen with bypass surgery, PTCA provides a useful alternative revascularization method for patients with less extensive disease, in whom the risk of surgery may equal or exceed any beneficial effect. New technology and growing experience are widening the scope of percutaneous revascularization by extending the hope of symptomatic relief and survival benefit even to patients with extensive, severe coronary artery disease. Comparisons between surgical therapy and PTCA in select populations with single- and multivessel coronary artery disease have shown that PTCA is not as effective as surgery for long-term symptomatic control, and that it often requires repeat PTCA or cross-over to bypass surgery; however, long-term outcomes (i.e., death and myocardial infarction) are similar. The cost of treatment beginning with PTCA may be lower than that of initial surgery, even when the increased need for repeat revascularization is taken into account. Despite this, surgical bypass remains the mainstay of therapy for patients with severe coronary artery disease and a poor prognosis for survival, and will remain the fallback procedure for patients who repeatedly undergo failed PTCA. At the present time, revascularization should be offered on the basis of symptom severity (in the presence of medical therapy) and in accordance with the prognosis for survival as judged by the extent and severity of disease (Table VI). Percutaneous transluminal coronary angioplasty is preferred in patients who require revascularization but can obtain no proven benefit from bypass surgery. Coronary artery bypass surgery, using the internal mammary artery when possible, remains the revascularization method of choice for patients with more severe disease or whose disease is not amenable to treatment using percutaneous methods (Table VII).

Angioplasty, Balloon, Coronary↗
PubMed1995

Gene therapy for human liver disease.

Investigators working in the area of gene therapy believe the potential for advances in all medical disciplines is enormous. It is humbling, however to appreciate how far we need to go, as the field is truly in its infancy. Gene transfer technologies currently under evaluation in clinical trials have major limitations. Vector systems used in the clinics by the year 2000 probably have yet to be discovered. An additional lesson learned is that efforts at gene therapy are hampered by a lack of knowledge of the basic biology of the target organ and pathogenesis of the underlying disease. Successful gene therapy programs will critically evaluate the field and through fundamental research move steadily forward toward the long-term goal of truly effective therapy for a wide spectrum of disease. In the near future, two liver diseases are the most likely to be treated with gene therapy. The evaluation of patients with familial hypercholesterolemia is ongoing, and once approved, more candidates will be enrolled for therapy. Progress also has been made in creating vectors for the treatment of ornithine transcarbamylase deficiency. It remains to be seen whether adenoviruses or retroviruses will be used first in attempts to control this disease. Although the inflammatory response noted with current recombinant viruses is a formidable problem, the efficiency of gene transfer into the liver with these vectors makes continued study worthwhile.

Adenoviridae↗
PubMed1994

Amygdala dopamine levels are markedly elevated after self- but not passive-administration of cocaine.

The influence of cocaine on rat brain monoaminergic neurotransmitters (dopamine, serotonin, noradrenaline) and their metabolites, and on binding of [3H]WIN 35,428 and [3H]GBR 12,935 to the dopamine transporter was measured after 4 weeks of cocaine exposure. Cocaine (mean daily dose 9.25 +/- 0.48 mg/kg) was self-administered (responders) or passively received (yoked) during sessions which lasted for 1 h per day. As compared with the controls, mean dopamine and serotonin levels were significantly elevated (+ 107% and + 47%, respectively) in amygdala of responders, but not of yoked rats, sacrificed 1 h after the last cocaine session. Dopamine and metabolite levels were normal in all other brain areas examined, including striatum, nucleus accumbens and medial prefrontal cortex, at both 1 h and 4 weeks withdrawal from cocaine. [3H]WIN 35,428 and [3H]GBR 12,935 binding were unaltered after cocaine exposure. These data provide additional support for the involvement of the amygdala in the acquisition of drug seeking behavior associated with cocaine self-administration.

Amygdala↗
PubMed1994

Inefficient gene transfer by adenovirus vector to cystic fibrosis airway epithelia of mice and humans.

The success of adenoviral vectors for gene therapy of lung disease in cystic fibrosis (CF) depends on efficient transfer of the complementary DNA encoding the correct version of the cystic fibrosis transmembrane regulator (CFTR) to the affected columnar epithelial cells lining the airways of the lung. Pre-clinical studies in vitro suggest that low doses of adenovirus vectors carrying this CFTR cDNA can correct defective Cl- transport in cultured human CF airway epithelia. Here we use mice carrying the disrupted CF gene to test the efficacy of this transfer system in vivo. We find that even repeated high doses can only partially (50%) correct the CF defect in Cl- transport in vivo and do not correct the Na+ transport defect at all. We investigated this discrepancy between the in vivo and in vitro transfer efficiency using CF mouse and human samples, and found that it reflects a difference in the susceptibility to adenovirus-5 transduction of the epithelial cell types dosed in vivo (columnar) and in vitro (basal-cell-like). These studies indicate that more efficient adenoviral gene-transfer vectors and/or refinement of dosing strategies are needed for therapy of CF lung disease.

Adenoviruses, Human↗
PubMed1994

Choline acetyltransferase activity is reduced in rat nucleus accumbens after unlimited access to self-administration of cocaine.

Choline acetyltransferase (ChAT) activity was measured in discrete areas of rat brain after chronic, unlimited access to self-administration of cocaine. Mean activity of ChAT was reduced by approximately 30% in the nucleus accumbens, both on the last day of cocaine access and after 3 weeks cocaine withdrawal. These data suggest that chronic cocaine exposure might inhibit nucleus accumbens cholinergic neurones which could underlie some of the behavioral effects of cocaine.

Animals↗
PubMed1994

Epidermal growth factor and transforming growth factor-alpha mRNA in rat small intestine: in situ hybridization study.

The expression of epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) mRNA in the small intestine of suckling and adult rats was examined by in situ hybridization. EGF mRNA was found mainly in the intestinal crypts in adult rats. Adult rats also exhibited a considerably stronger signal for EGF mRNA in comparison to suckling rats, where the signal was very low or absent. In contrast to EGF, very strong expression of TGF-alpha mRNA was observed in the small intestine of both adult and suckling rats. These data suggest the differences between the expression of EGF and TGF-alpha in the developing small intestine.

Aging↗
PubMed1994

Ablation of E2A in recombinant adenoviruses improves transgene persistence and decreases inflammatory response in mouse liver.

First-generation recombinant adenoviruses that lack E1 sequences have shown tremendous promise in animal and human models of gene therapy. Important limitations of these vectors are that recombinant gene expression is transient and inflammation occurs at the site of gene transfer. Our hypothesis for generating vectors with increased persistence is that present recombinant adenoviruses express viral proteins that stimulate cellular immune responses leading to destruction of the infected cells and repopulation of the organ with non-transgene-containing cells. This model predicts that further crippling of the virus will improve persistence and diminish pathology. We describe in this report second-generation recombinant adenoviruses harboring a beta-galactosidase-expressing transgene in which a temperature-sensitive mutation has been introduced into the E2A gene of an E1-deleted recombinant. At nonpermissive temperature, this virus fails to express late gene products, even when E1 is expressed in trans. The biology of this recombinant was studied in vivo in the context of mouse liver, a setting that is permissive for adenovirus type 5 replication. Animals that received the second-generation virus expressed the transgene for at least 70 days, whereas expression of the first-generation virus was no longer than 14 days. In addition, the inflammatory response, as measured by infiltration of CD8+ T cells, was blunted and delayed in livers infected with second-generation virus. These studies illustrate that modifications that disrupt structural protein expression in recombinant adenoviruses may be useful in enhancing their utility for gene therapy.

Adenoviridae↗
PubMed1994

Cellular immunity to viral antigens limits E1-deleted adenoviruses for gene therapy.

An important limitation that has emerged in the use of adenoviruses for gene therapy has been loss of recombinant gene expression that occurs concurrent with the development of pathology in the organ expressing the transgene. We have used liver-directed approaches to gene therapy in mice to study mechanisms that underlie the problems with transient expression and pathology that have characterized in vivo applications of first-generation recombinant adenoviruses (i.e., those deleted of E1a and E1b). Our data are consistent with the following hypothesis. Cells harboring the recombinant viral genome express the transgene as desired; however, low-level expression of viral genes also occurs. A virus-specific cellular immune response is stimulated that leads to destruction of the genetically modified hepatocytes, massive hepatitis, and repopulation of the liver with nontransgene-containing hepatocytes. These findings suggest approaches for improving recombinant adenoviruses that are based on further crippling the virus to limit expression of nondeleted viral genes.

Adenovirus E1A Proteins↗
PubMed1994

In vivo correction of low density lipoprotein receptor deficiency in the Watanabe heritable hyperlipidemic rabbit with recombinant adenoviruses.

A rabbit animal model of the human disease familial hypercholesterolemia (FH), which is the result of low density lipoprotein (LDL) receptor deficiency, was used to develop an in vivo approach to gene therapy based on recombinant adenoviruses. Recombinant, replication-defective adenoviruses expressing the lacZ gene under the control of different promoters were infused into the portal circulation of New Zealand White (NZW) rabbits. Expression of lacZ could be obtained in virtually all hepatocytes within 3 days post-infusion, but was undetectable by 3 weeks. This was not associated with liver pathology. An LDL receptor expressing adenovirus was constructed using the most active promoter and was infused into the portal vein of rabbits deficient in LDL receptor. Analysis of liver tissues harvested 3 days after virus infusion demonstrated human LDL receptor protein in the majority of hepatocytes that exceeded the levels found in human liver by at least 10-fold. Transgene expression was stable for 7-10 days and diminished to undetectable levels within 3 weeks. Infusion of LDL receptor expressing virus led to substantial reductions in serum cholesterol that returned to base line within 3 weeks; this acute reduction in serum cholesterol was associated with accumulations of lipid in hepatocytes. The development of neutralizing antibodies to the recombinant adenovirus markedly diminished the effectiveness of a second dose. These studies illustrate the advantages of recombinant adenoviruses for the treatment of liver metabolic diseases and define issues, such as viral genome instability and blocking immune response, that need to be overcome before the promise of this technology can be fully realized.

Adenoviruses, Human↗
PubMed1994

Use of recombinant adenovirus to transfer the herpes simplex virus thymidine kinase (HSVtk) gene to thoracic neoplasms: an effective in vitro drug sensitization system.

Transfer of the herpes simplex virus thymidine kinase (HSVtk) gene into tumor cells using retroviral vectors followed by administration of ganciclovir provides a potential strategy for the treatment of malignancy. Because of the limitations of using retroviral vectors for clinical application, the feasibility of using a recombinant adenovirus containing HSVtk was examined. Cell lines derived from human malignant mesotheliomas and non-small cell lung cancers infected with a recombinant adenovirus containing HSVtk showed strong expression of HSVtk protein as determined by immunohistochemical staining. Infection with a recombinant adenovirus containing HSVtk rendered cells sensitive to doses of ganciclovir that were 2-3 logs lower than uninfected cells or those infected with a control virus. A strong "bystander effect" was noted in mesothelioma lines; there was no diminution in the efficacy of ganciclovir treatment until the ratio of infected:uninfected cells fell below 1:10. This study thus demonstrates in vitro efficacy of an adenovirus-transduced HSVtk drug sensitization gene therapy system in thoracic malignancies. Recombinant adenovirus transfer of the HSVtk gene followed by ganciclovir may have promise as an in situ treatment for tumors.

Adenoviridae↗