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Biomedical subjects

E Holland

Publications and source records attributed to E Holland.

At least 19 recordsLinked to original sources
PubMed2002

TEL Induces Aggregation in Transformed Cells and Induces Tube Formation in NIH3T3-UCLA Cells.

TEL/ETV6 is the frequent target of translocations associated with lymphoid and myeloid leukemias and solid tumors. We show that TEL induces aggregation of immortalized and transformed fibroblasts, endothelial cells and astrocytes. These aggregates form cellular cords in NIH3T3-UCLA by a cell autonomous process, which occurs when the monolayer is made up of over 75% of cells expressing exogenous TEL. Cords with a diameter of 15-25 microm contain a lumen and occur as tube structures. The possible relevance for vasculogenic mimicry is discussed. By contrast TEL did not induce aggregation of regular NIH3T3 cells, an effect that could only be induced by co-expression of oncogenic RAS/Lys12. Also transduction of TEL and RAS retroviral vectors into the endothelial MS1 cell line and TEL alone in the highly transformed glioblastoma cell lines EH-A and EH-B resulted in extensive aggregation. Thus, the induction of cellular aggregation by TEL correlates with transformation.

3T3 Cells↗
PubMed2001

Consistent land- and atmosphere-based U.S. carbon sink estimates.

For the period 1980-89, we estimate a carbon sink in the coterminous United States between 0.30 and 0.58 petagrams of carbon per year (petagrams of carbon = 10(15) grams of carbon). The net carbon flux from the atmosphere to the land was higher, 0.37 to 0.71 petagrams of carbon per year, because a net flux of 0.07 to 0.13 petagrams of carbon per year was exported by rivers and commerce and returned to the atmosphere elsewhere. These land-based estimates are larger than those from previous studies (0.08 to 0.35 petagrams of carbon per year) because of the inclusion of additional processes and revised estimates of some component fluxes. Although component estimates are uncertain, about one-half of the total is outside the forest sector. We also estimated the sink using atmospheric models and the atmospheric concentration of carbon dioxide (the tracer-transport inversion method). The range of results from the atmosphere-based inversions contains the land-based estimates. Atmosphere- and land-based estimates are thus consistent, within the large ranges of uncertainty for both methods. Atmosphere-based results for 1980-89 are similar to those for 1985-89 and 1990-94, indicating a relatively stable U.S. sink throughout the period.

Agriculture↗
PubMed2001

Eye banking and screening for Creutzfeldt-Jakob disease.

OBJECTIVES: To quantify the risk of Creutzfeldt-Jakob disease (CJD) among cornea donors, evaluate supplemental screening strategies, and address concerns about the adequacy of current methods of screening tissue donors in the United States. METHODS: Reported data on deaths due to CJD and from all causes were used to estimate the rate of CJD among cornea donors. The impact of increased screening on risk of CJD and donor supply was evaluated. RESULTS: Only 1.3 of the approximately 45 000 cornea donors in the United States each year might be expected to have CJD. Most of the estimated risk (91%) is due to preclinical (asymptomatic) disease and therefore could not be eliminated by screening for signs or symptoms. If only the highest-risk age group (60 to 69 years) were screened and specificity were 90%, more than 21 000 otherwise acceptable donors would incorrectly be excluded over a period of 17.5 years to correctly exclude a single donor with symptomatic CJD. CONCLUSIONS: Currently, the risk of CJD transmission following cornea transplantation is remarkably low. Screening for symptoms of CJD would have minimal impact on safety, but would reduce donor supply and likely result in many patients not receiving needed treatment.

Adolescent↗
PubMed2001

Human airway smooth muscle cells secrete vascular endothelial growth factor: up-regulation by bradykinin via a protein kinase C and prostanoid-dependent mechanism.

Bronchial vascular remodeling is an important feature of the pathology of chronic asthma, but the responsible mechanisms and main sources of angiogenic factors are unclear. Here we report that human airway smooth muscle cells express vascular endothelial growth factor (VEGF)121, 165, 189, 206 splice variants and secrete VEGF protein constitutively. VEGF protein secretion was increased by the proinflammatory asthma mediator bradykinin through post-transcriptional mechanisms. Bradykinin-induced VEGF secretion was dependent on the B2 bradykinin receptor, activation of protein kinase C, and generation of endogenous prostanoids. This is the first report that bradykinin can increase VEGF secretion in any biological system and the first to show that airway smooth muscle cells produce VEGF. Our results suggest a novel role for human airway smooth muscle in contributing to bronchial mucosal angiogenesis in chronic asthma by secretion of VEGF and suggest a wider role for mesenchymal cell products in mediating angiogenesis in inflammatory and allergic diseases.

Adult↗
PubMed2001

A multivariate analysis of 416 patients with glioblastoma multiforme: prognosis, extent of resection, and survival.

OBJECT: The extent of tumor resection that should be undertaken in patients with glioblastoma multiforme (GBM) remains controversial. The purpose of this study was to identify significant independent predictors of survival in these patients and to determine whether the extent of resection was associated with increased survival time. METHODS: The authors retrospectively analyzed 416 consecutive patients with histologically proven GBM who underwent tumor resection at the authors' institution between June 1993 and June 1999. Volumetric data and other tumor characteristics identified on magnetic resonance (MR) imaging were collected prospectively. CONCLUSIONS: Five independent predictors of survival were identified: age, Karnofsky Performance Scale (KPS) score, extent of resection, and the degree of necrosis and enhancement on preoperative MR imaging studies. A significant survival advantage was associated with resection of 98% or more of the tumor volume (median survival 13 months, 95% confidence interval [CI] 11.4-14.6 months), compared with 8.8 months (95% CI 7.4-10.2 months; p < 0.0001) for resections of less than 98%. Using an outcome scale ranging from 0 to 5 based on age, KPS score, and tumor necrosis on MR imaging, we observed significantly longer survival in patients with lower scores (1-3) who underwent aggressive resections, and a trend toward slightly longer survival was found in patients with higher scores (4-5). Gross-total tumor resection is associated with longer survival in patients with GBM, especially when other predictive variables are favorable.

Adult↗
PubMed2000

Selectivity of cyclo-oxygenase inhibitors in human pulmonary epithelial and smooth muscle cells.

Cyclo-oxygenase (COX) inhibitors may have a role in reducing inflammation in asthma and other pulmonary diseases. COX inhibitors have different selectivities for the two COX isoenzymes (COX 1 and COX 2) which vary between purified enzyme and intact cell preparations. The relative selectivity of COX inhibitors has not been studied in human airway cells. A number of COX inhibitors in cultured human airway cells were compared which exclusively express either COX 1 (primary degree cultured human airway smooth muscle (HASM) cells) or COX 2 (A549 pulmonary epithelial cell-line) as measured by Western blotting. COX activity was assayed by prostaglandin (PG)E2 production following 30 min incubation with 5 mM arachidonic acid. COX activity in both cell types was similar; HASM cells 92.2+/-12.1 ng PGE2 x mg-1 protein, A549 cells 87.7+/-24.4 ng PGE2 mg-1 protein. In HASM cells the median inhibitory concentration (IC50) was >10-5 M for nimesulide, 3.2 x 10-6 M for N-(2-cyclohexyloxy-4-nitrophenyl)-methanesulphonamide (NS398), 1.8 x 10-8 M for flurbiprofen, 6.7 x 10-9 M for indomethacin and >10-5 M for aspirin. In A549 cells the IC50 was 1.8 x 10-9M for nimesulide, 4.1 x 10-9 M for NS398,6.2 x 10-10 M for flurbiprofen, 1.3 x 10-8 M for indomethacin and >10-5 M for aspirin. Sodium valerate had no effect in either HASM or A549 cells. The COX 2:COX 1 selectivity ratio (COX 2 IC50/COX I IC50) was <0.0001 for nimesulide, 0.001 for NS398, 0.03 for flurbiprofen and 1.9 for indomethacin. In conclusion the present study has shown that cyclo-oxygenase inhibitors have a range of selectivities for cyclo-oxygenase 1 and cyclo-oxygenase 2 in intact human airway cells. The relative cyclo-oxygenase 2 selectivity of N-(2-cyclohexyloxy-4-nitrophenyl)-methanesulphonamide and nimesulide may have implications for the treatment of asthma and other inflammatory pulmonary diseases.

Adult↗
PubMed2000

TGF-beta1 stimulates IL-8 release, COX-2 expression, and PGE(2) release in human airway smooth muscle cells.

We have recently shown that endogenous prostanoids are critical in bradykinin-stimulated interleukin (IL)-8 release from human airway smooth muscle (ASM) cells. In this study, we tested the ability of transforming growth factor (TGF)-beta1 to stimulate IL-8 release, cyclooxygenase (COX)-2 expression and PGE(2) generation in cultured human ASM cells and explored the role of COX products and COX-2 induction on IL-8 release. TGF-beta1 stimulated IL-8 release, COX-2 induction, and PGE(2) generation in a concentration- and time-dependent manner. Maximal IL-8 release was achieved with 10 ng/ml of TGF-beta1 after 16 h of incubation, which was inhibited by the transcription inhibitor actinomycin D and the corticosteroid dexamethasone but was not affected by the nonselective COX inhibitor indomethacin and the selective COX-2 inhibitor NS-398 despite their inhibition on TGF-beta1-induced PGE(2) release. These results show for the first time that TGF-beta1 stimulates IL-8 release, COX-2 induction, and PGE(2) generation in human ASM cells and that PGE(2) generation is not critical for TGF-beta1-induced IL-8 release. These findings suggest that TGF-beta1 may play an important role in the pathophysiology of asthma.

Aged↗
PubMed1999

Development of a flexible and specific gene delivery system for production of murine tumor models.

To develop models of human cancer we have expressed the avian retroviral receptor, TVA, under a variety of mammalian promoters in transgenic mice, thus rendering mice susceptible to infection with avian leukosis virus-derived gene vectors. TVA-based retroviral gene transfer offers advantages over current murine models of human cancer. A single transgenic mouse line can be used to evaluate multiple genetic lesions, individually and in combination. Furthermore, mutant genes are introduced somatically into animals, as occurs in the majority of naturally occurring tumors. Because the avian viral vectors replicate only in avian cells, the viral receptor in infected transgenic mouse cells remains available for multiple rounds of infection with different ASLV vectors. We discuss the theoretical and practical aspects of using recombinant avian retroviruses with TVA transgenic mice to generate cancer models.

Alpharetrovirus↗
PubMed1999

Immunocytochemical localization of cyclo-oxygenase isoforms in cultured human airway structural cells.

BACKGROUND: Cyclo-oxygenase (COX) exists as two isoforms, COX-1, the constitutive isoform, and COX-2, which is inducible by cytokines or inflammatory stimuli and may participate in airway inflammation. OBJECTIVE: To determine the basal distribution of COX isoforms, and their regulation by interleukin-1 beta (IL-1beta), bradykinin (BK) and dexamethasone (Dex) in cultured airway structural cells. METHODS: We measured COX-1 and COX-2 in cultured human airway smooth muscle (HASM) cells, MRC5 fibroblasts and normal human epithelial cells (NHBE) using immunocytochemical analysis. RESULTS: The majority of all types of untreated cultured cells expressed COX-1 (75% of HASM, 75% of MRC5 fibroblasts and 72% of NHBE cells). Fibroblasts and smooth muscle cells showed low constitutive COX-2 expression (2 and 8%, respectively) but this was higher in NHBE cells (28%). IL-1beta (24 h incubation) or BK (4 h incubation) had no effect on COX-1 expression in any of the cells studied. In contrast, there was a two- and 1.5-fold rise in the percentage of NHBE cells expressing COX-2; a 7.5- and sixfold rise in the percentage of HASM cells expressing COX-2 and a 33.5- and 20.5-fold increase in the percentage of fibroblasts expressing COX-2 after IL-1beta or BK treatment, respectively. Pretreatment with dexamethasone abolished IL-1beta- and BK-stimulated COX-2 induction in all cells studied. CONCLUSION: COX-1 is expressed constitutively in human airway fibroblasts, smooth muscle and epithelial cells but epithelial cells also show constitutive expression of COX-2. Both IL-1beta and BK induced COX-2 expression in all cells studied and this induction was blocked by dexamethasone. Immunocytochemical techniques can be successfully used to detect the distribution of COX isoforms in cell cultures.

Blotting, Western↗
PubMed1999

Desensitization of guanylyl cyclases in cultured human airway smooth-muscle cells.

We previously showed that cultured human airway smooth-muscle cells (HASMC) contain soluble and particulate guanylyl cyclases (GCs). We studied the desensitization of soluble and particulate GCs in HASMC. Homologous desensitization of soluble GC occurred after incubation with S-nitroso-N-acetyl pencillamine (SNAP). SNAP-dependent desensitization was blocked by hemoglobin, a nitric oxide (NO) scavenger, suggesting that it was due to NO release. Cross-desensitization between SNAP and sodium nitroprusside (SNP) and the lack of thiol reduction after SNAP or SNP treatment suggested that thiol depletion was not involved. Assays for soluble GC activity and experiments using protein synthesis inhibitors suggested that SNAP-dependent desensitization was due to reduced soluble GC. Homologous desensitization of particulate GC occurred after pretreatment with atrial natriuretic peptide (ANP) accompanied by reduced particulate GC activity. Recovery required protein synthesis, suggesting that it was due to reduction in particulate GC. Homologous desensitization to either SNAP or ANP was not altered by phosphodiesterase (PDE) inhibitors, suggesting that increased PDE activity was not involved. Cross-desensitization experiments using SNAP and ANP and experiments using zaprinast to elevate cyclic guanosine monophosphate showed no evidence of heterologous desensitization. Our results suggest that pretreatment of HASMC with SNAP or ANP causes homologous, but not heterologous, desensitization of soluble and particulate GCs, respectively.

Atrial Natriuretic Factor↗
PubMed1998

Role of cyclo-oxygenase-2 induction in interleukin-1beta induced attenuation of cultured human airway smooth muscle cell cyclic AMP generation in response to isoprenaline.

Airway smooth muscle (ASM) in human asthma shows reduced relaxation and cyclic AMP generation in response to beta-adrenoceptor agonists. IL-beta attenuates cyclic AMP generation but the underlying mechanism is unclear. We have reported that IL-1beta induces cyclo-oxygenase-2 (COX-2) in human ASM cells and results in a marked increase in prostanoid generation with PGE2 and PGI2 as the major products. We investigated the role of COX-2 induction and prostanoid release (measured as PGE2) in IL-1beta induced attenuation of cyclic AMP generation in response to the beta-adrenoceptor agonist isoprenaline (ISO). Pre-treatment of human ASM cells with IL-1beta significantly attenuated cyclic AMP generation in response to high concentrations of ISO (1.0-10.0 microM) in a time- and concentration-dependent manner. The effect was accompanied by a high concentration of PGE2 release. The non-selective COX inhibitor indomethacin (Ind), the selective COX-2 inhibitor NS-398, the protein synthesis inhibitors cycloheximide (CHX) and actinomycin D and the steroid dexamethasone (Dex) all abolished the PGE2 release and prevented the attenuated cyclic AMP generation. COX substrate arachidonic acid time- and concentration-dependently mimicked IL-1beta induced attenuation and the effect was prevented by the non-selective COX inhibitors Ind and flurbiprofen, but not by NS-398, CHX and Dex. In contrast to IL-1beta, TNFalpha and IFNgamma, which are ineffective in inducing COX-2 and releasing PGE2 from human ASM cells, did not affect the cyclic AMP formation. Our study demonstrates that COX-2 induction and the consequent release of prostanoids plays a crucial role in IL-1beta induced attenuation of human ASM cell cyclic AMP response to ISO.

Adrenergic beta-Agonists↗
PubMed1998

Impaired cAMP production in human airway smooth muscle cells by bradykinin: role of cyclooxygenase products.

Interleukin (IL)-1beta impairs human airway smooth muscle (ASM) cell cAMP responses to isoproterenol (Iso). We investigated if bradykinin (BK) could cause a similar effect and the role of cyclooxygenase (COX) products in this event, since we have recently reported that BK, like IL-1beta, also causes COX-2 induction and prostanoid release in human ASM cells. BK pretreatment significantly attenuated Iso-induced cAMP generation in a time- and concentration-dependent manner. cAMP generation by prostaglandin (PG) E2 but not by forskolin was also impaired. The COX inhibitor indomethacin completely prevented the impairment, whereas the selective COX-2 inhibitors NS-398 and nimesulide, protein synthesis inhibitors cycloheximide and actinomycin D, and steroid dexamethasone were all partially effective. The impairment was mimicked by the B2 agonist [Tyr(Me)8]BK, the Ca2+ ionophore A-23187, and PGE2 and prevented by the B2 antagonist HOE-140, but anti-IL-1beta serum was ineffective. The results indicate that BK impairs human ASM cell responses to Iso, and the effect is largely mediated by B2 receptor-related COX product release via both COX isoforms and is independent of IL-1beta.

Adult↗
PubMed1997

Na+/K+ ATPase in lower airway epithelium from cystic fibrosis and non-cystic-fibrosis lung.

The basolateral Na+/K+ ATPase plays a critical role in sodium reabsorption across airway epithelium. Nasal epithelium shows increased Na+/K+ ATPase activity in cystic fibrosis (CF) but Na+/K+ ATPase has not been characterized in human lung epithelium or compared in CF and normal lung. We measured 3[H] ouabain binding and Na+/K+ ATPase activity in human tracheal epithelium and compared Na/K ATPase activity in bronchial epithelium in CF and control subjects. In tracheal epithelium Na+/K+ ATPase pumps were abundant and of high 3[H] ouabain affinity (Kd 4.7 nM, Bmax 38 pmol/mg) and Na+/K+ ATPase activity was 55 +/- 8 nmol/mg protein/min. Bronchial epithelial Na+/K+ ATPase activity was twofold higher in CF patients than in controls. The increased Na+/K+ ATPase activity may contribute to the increased sodium reabsorption seen in cystic fibrosis.

Adolescent↗
PubMed1997

Regulation of cGMP by soluble and particulate guanylyl cyclases in cultured human airway smooth muscle.

Although guanosine 3',5'-cyclic monophosphate (cGMP) acts as a relaxant second messenger, the regulation of intracellular cGMP has not been comprehensively studied in human airway smooth muscle. We studied the production of cGMP by cultured human airway smooth muscle cells (HASMC) after stimulation with activators of soluble guanylyl cyclase [sodium nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine (SNAP)] and particulate guanylyl cyclase [atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), C-type natriuretic peptide (CNP), and Escherichia coli heat stable enterotoxin (STa)]. cGMP was measured by enzyme-linked immunosorbent assay. Both SNP (10(-6) to 10(-3) M) and SNAP (10(-6) to 10(-3) M) caused concentration-dependent elevation of cGMP in the presence of the nonselective phosphodiesterase (PDE) inhibitor 3-isobutyl-1-methylxanthine (10(-3) M), with cGMP increasing 6- and 15-fold in response to SNP and SNAP, respectively, at the highest concentration tested (10(-3) M). The increases in cGMP in response to SNP (5 x 10(-5) M) and SNAP (10(-5) M) were inhibited by hemoglobin (Hb; 5 x 10(-5) M), a nitric oxide scavenger, and methylene blue (MB; 5 x 10(-4) M), an inhibitor of guanylyl cyclase. cGMP accumulation after SNAP was abolished by both Hb and MB. The response to SNP was inhibited by 79% with Hb and was abolished with MB. ANP, BNP, and CNP (10(-9) to 10(-5) M) + phosphoramidon (10(-6) M) caused a concentration-dependent elevation in cGMP with an order of potency ANP > BNP > CNP. cGMP formation in the presence of the highest concentration of the most potent natriuretic peptide (10(-5) M ANP) was two- to threefold greater than with the highest concentration of SNAP. The increase in cGMP seen with natriuretic peptides was similar in the presence or absence of phosphoramidon, a neutral endopeptidase (NEP) inhibitor, suggesting that NEP is not playing a role in modulating the effect of natriuretic peptides in HASMC. STa (400 IU/ml) had no effect on cGMP levels. SNAP- and ANP-induced cGMP accumulation was increased by the selective type V PDE inhibitors SKF-96231 and zaprinast, suggesting that type V PDE is responsible for cGMP breakdown in HASMC. These results suggest that cultured HASMC contain both soluble and particulate guanylyl cyclases. The order of potency of the natriuretic peptides ANP > BNP > CNP suggests that type A particulate membrane-bound guanylate cyclase predominates.

1-Methyl-3-isobutylxanthine↗
PubMed1996

Diagnosis of microsporidial keratitis by confocal microscopy and the chromatrope stain.

PURPOSE: To illustrate the value of confocal microscopy and chromatrope stain in the diagnosis of microsporidial keratitis. METHODS: In vivo confocal microscopy was performed on a man with the human immunodeficiency virus who had severe bilateral epithelial keratitis refractory to topical antibacterial medications. The results were compared to conjunctival scrapings stained with the chromatrope-based Weber stain. RESULTS: Confocal microscopy demonstrated many small, intraepithelial opacities of the corneal epithelium, which were suggestive of Microsporidia. Results of the chromatrope stain of conjunctival scrapings confirmed the diagnosis of microsporidial keratitis. CONCLUSIONS: Rapid diagnosis allowed prompt initiation of topical fumagillin, which permitted rapid, long-term control of the symptoms of microsporidial keratitis.

AIDS-Related Opportunistic Infections↗
PubMed1995

Production of PGE2 by bovine cultured airway smooth muscle cells: regulation by cAMP.

Prostaglandin E2 (PGE2) is thought to be an important inhibitory modulator of inflammatory processes in the airway. Previous studies have shown that it is produced by bovine cultured airway smooth muscle (ASM) cells in large quantities, but its regulation by second messengers has not been studied in this tissue. To determine whether PGE2 production by ASM might be an important action of beta-adrenoceptor agonists in asthma, the regulation of PGE2 production by adenosine 3',5'-cyclic monophosphate (cAMP) was assessed using dibutyryl cAMP (DBcAMP), forskolin, and albuterol. DBcAMP increased PGE2 production over a 24-h time course. Forskolin and albuterol both increased PGE2 production over control cells to similar levels after 24 h. Incubation of albuterol-treated cells with propranolol significantly (70%) reduced the stimulatory effect of albuterol on PGE2 production. Incubation of forskolin-treated cells with Rp-cAMP, a cAMP antagonist, inhibited the PGE2 response evoked by forskolin by 80%. Ro-20-1724, a selective inhibitor of type IV phosphodiesterase, stimulated PGE2 production (P = 0.02). Cycloheximide, a protein-synthesis inhibitor, did not inhibit the response to DBcAMP. The effects of DBcAMP were additive with the effects of bradykinin, a proinflammatory mediator known to increase PGE2 production (P < 0.05). These studies suggest that cAMP may play an important regulatory role in stimulating PGE2 production by ASM. This may be a novel beneficial action of beta-adrenoceptor agonists in asthma.

4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone↗