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J M Routes

Publications and source records attributed to J M Routes.

At least 19 recordsLinked to original sources
PubMed2001

MHC class I molecules on adenovirus E1A-expressing tumor cells inhibit NK cell killing but not NK cell-mediated tumor rejection.

Expression of adenovirus E1A gene products in tumor cells enhances NK cell lysis in vitro and NK-mediated rejection in vivo, despite increasing class I molecules on tumor cells. It is unclear why the increased expression of MHC class I molecules does not appear to confer resistance to killing by NK cells. One possibility is the unique capacity of E1A to sensitize cells to multiple NK cell killing mechanisms including perforin/granzyme, Fas ligand, tumor necrosis factor-alpha and TRAIL. To examine this issue, MCA-102-E1A tumor cells (H-2(b)) that express E1A and are NK sensitive were transfected with H-2D(d), the ligand for the NK inhibitory receptor, Ly49A. Expression of H-2D(d) molecules by MCA-102-E1A cells protected them from lysis by a Ly49A(+) NK cell clone and Ly49A(+) NK cells isolated from C57BL/6 nude mice. In contrast, NK cell-mediated rejection of MCA-102-E1A tumor cells was not inhibited by the expression of H-2D(d) molecules, nor was killing by polyclonal populations of NK cells isolated from C57BL/6-nude mice. H-2D(d) interacts with several inhibitory Ly49 receptors that are non-clonally expressed on NK cells in C57BL/6 mice: Ly49A (20% of NK cells), Ly49G2 (54% of NK cells) and Ly49C/I (47% of NK cells). Our data indicate that while E1A sensitizes cells to NK cell killing, it does not interfere with signal transduction by inhibitory NK receptors. Therefore, a small population of NK cells that do not express Ly49A, Ly49G2 or Ly49C/I inhibitory receptors are likely responsible for the rejection of MCA-102-E1A-D(d) tumor cells in vivo.

Adenovirus E1A Proteins↗
PubMed2001

Role of the innate immune response in determining the tumorigenicity of neoplastic cells.

Expression of the E1A oncogene of adenovirus serotype 5 (Ad5) sensitizes neoplastic cells from different species, including human, to apoptosis triggered by components of the innate mmune response. This E1A activity causes a marked reduction in the tumorigenicities of sarcoma cells in a manner that is directly linked to the competence of NK cells from the test animals to lyse E1A-expressing tumour cells. The results from this oncogene model indicate that studies of quantitative differences in the tumorigenicities of different neoplastic cells should include considerations of oncogene regulation of susceptibility to immune-mediated injury and the immunocompetence of the animals chosen as tumour challenge recipients.

Animals↗
PubMed2000

CREB (cAMP response element binding protein) and C/EBPalpha (CCAAT/enhancer binding protein) are required for the superstimulation of phosphoenolpyruvate carboxykinase gene transcription by adenoviral E1a and cAMP.

In the present study, we observed superstimulated levels of cAMP-stimulated transcription from the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter in cells infected with wild-type adenovirus expressing 12 S and 13 S E1a proteins, or in cells expressing 13 S E1a alone. cAMP-stimulated transcription was inhibited in cells expressing only 12 S E1a, but slightly elevated in cells expressing E1a proteins with mutations in conserved regions 1 or 2, leading us to conclude that the superstimulation was mediated by conserved region 3 of 13 S E1a. E1a failed to enhance cAMP-stimulated transcription from promoters containing mutations that abolish binding by cAMP response element binding protein (CREB) or CCAAT/enhancer binding proteins (C/EBPs). This result was supported by experiments in which expression of dominant-negative CREB and/or C/EBP proteins repressed E1a- and cAMP-stimulated transcription from the PEPCK gene promoter. In reconstitution experiments using a Gal4-responsive promoter, E1a enhanced cAMP-stimulated transcription when chimaeric Gal4-CREB and Gal4-C/EBPalpha were co-expressed. Phosphorylation of CREB on serine-133 was stimulated in cells treated with dibutyryl cAMP, whereas phosphorylation of C/EBPalpha was increased by E1a expression. Our data support a model in which cAMP agonists increase CREB activity and stimulate PEPCK gene transcription, a process that is enhanced by E1a through the phosphorylation of C/EBPalpha.

Adenovirus E1A Proteins↗
PubMed2000

Dissimilar immunogenicities of human papillomavirus E7 and adenovirus E1A proteins influence primary tumor development.

Although human papillomaviruses (HPV) and adenoviruses (Ad) both transform cells by expressing functionally related oncogenes (Ad-E1A/E1B; HPV-E7/E6), only HPV are oncogenic in humans. Prior studies have shown that HPV-transformed cells are resistant to NK cell lysis and E7- and E6-specific CTL are inefficiently generated in women with HPV-induced cervical cancer. Therefore, we postulated that the dissimilar oncogenicities of Ad and HPV may be caused by a protective NK and T cell response that is triggered by transformed cells expressing E1A, but not by E7. To test this hypothesis, mice that were either immunologically intact, lacked T cells, or lacked both NK and T cells were challenged with Ad serotype 5 (Ad5)-E1A- or HPV16-E7-transfected tumor cells. E7-expressing tumor cells were resistant to NK cell lysis in vitro and failed to elicit a measurable anti-tumor NK or T cell response in vivo. The concomitant expression of E6 did not change this phenotype. In contrast, E1A-expressing tumor cells were sensitive to NK lysis in vitro and triggered a protective NK and T cell immune response in vivo. These data suggest differences in the capacities of E1A or E7 oncoproteins to trigger protective anti-tumor immune responses may contribute to the dissimilar oncogenicities of Ad and HPV in humans.

Adenovirus E1A Proteins↗
PubMed2000

Adenovirus E1A oncogene expression in tumor cells enhances killing by TNF-related apoptosis-inducing ligand (TRAIL).

Expression of the adenovirus serotype 5 (Ad5) E1A oncogene sensitizes cells to apoptosis by TNF-alpha and Fas-ligand. Because TNF-related apoptosis-inducing ligand (TRAIL) kills cells in a similar manner as TNF-alpha and Fas ligand, we asked whether E1A expression might sensitize cells to lysis by TRAIL. To test this hypothesis, we examined TRAIL-induced killing of human melanoma (A2058) or fibrosarcoma (H4) cells that expressed E1A following either infection with Ad5 or stable transfection with Ad5-E1A. E1A-transfected A2058 (A2058-E1A) or H4 (H4-E1A) cells were highly sensitive to TRAIL-induced killing, but Ad5-infected cells expressing equally high levels of E1A protein remained resistant to TRAIL. Infection of A2058-E1A cells with Ad5 reduced their sensitivity to TRAIL-dependent killing. Therefore, viral gene products expressed following infection with Ad5 inhibited the sensitivity to TRAIL-induced killing conferred by transfection with E1A. E1B and E3 gene products have been shown to inhibit TNF-alpha- and Fas-dependent killing. The effect of these gene products on TRAIL-dependent killing was examined by using Ad5-mutants that did not express either the E3 (H5dl327) or E1B-19K (H5dl250) coding regions. A2058 cells infected with H5dl327 were susceptible to TRAIL-dependent killing. Furthermore, TRAIL-dependent killing of A2058-E1A cells was not inhibited by infection with H5dl327. Infection with H5dl250 sensitized A2058 cells to TRAIL-induced killing, but considerably less than H5dl327-infection. In summary, expression of Ad5-E1A gene products sensitizes cells to TRAIL-dependent killing, whereas E3 gene products, and to a lesser extent E1B-19K, inhibit this effect.

Adenovirus E1A Proteins↗
PubMed1999

E1A oncogene induction of cellular susceptibility to killing by cytolytic lymphocytes through target cell sensitization to apoptotic injury.

E1A oncogene expression increases mammalian cell susceptibility to lysis by cytolytic lymphocytes (CLs) at a stage in this intercellular interaction that is independent of cell surface recognition events. Since CLs can induce either apoptotic or necrotic cell death, we asked whether E1A sensitization to injury-induced apoptosis is sufficient to explain E1A-induced cytolytic susceptibility. Mouse, rat, hamster, and human cells that were rendered cytolytic susceptible by E1A were also sensitized to CL-induced and chemically induced apoptosis. In contrast, E1A-positive cells were no more susceptible to injury-induced necrosis than E1A-negative cells. Similar to induction of cytolytic susceptibility and in contrast to other E1A activities, cellular sensitization to chemically induced apoptosis depended on high-level E1A oncoprotein expression. Loss of both cytolytic susceptibility and sensitization to chemically induced apoptosis was coselected during in vivo selection of E1A-positive sarcoma cells for increased tumorigenicity. Furthermore, E1A mutant proteins that cannot bind the cellular transcriptional coactivator, p300, and that fail to induce cytolytic susceptibility also failed to sensitize cells to injury-induced apoptosis. These data indicate that E1A induces susceptibility to killer cell-induced lysis through sensitization of cells to injury-induced apoptosis.

3T3 Cells↗
PubMed1999

Noninfectious immunologic disorders in adults.

This article covers a variety of immunologic defects and discusses their potential side effects, with an emphasis on immunodeficiencies with survival into adulthood. To organize the approach to immunodeficiencies, the immune system is broken down broadly into four major components: the complement cascade, antibody (B cell)-mediated immunity, cellular (T cell)-mediated immunity, and phagocytosis.

Adult↗
PubMed1996

Adenovirus E1A proteins regulate phosphoenolpyruvate carboxykinase gene transcription through multiple mechanisms.

Recently, Kalvakolanu et al. (Kalvakolanu, D. V. R., Liu, J., Hanson, R. W., Harter, M. L., and Sen, G. C. (1992) J. Biol. Chem. 267, 2530-2536) showed that E1A inhibited the basal and cAMP-stimulated transcription of the gene for phosphoenolpyruvate carboxykinase (PEPCK). This inhibition was mediated by the conserved region 1 (CR1) domain of E1A, which has been shown by other laboratories to bind to the cellular transcriptional adaptor proteins, p300 and cAMP response element binding protein (CREB)-binding protein (CBP). The PEPCK gene promoter contains a functional cAMP-response element, through which CREB and, therefore, CBP modulate transcription, and a consensus p300 DNA binding sequence is also present in a distal protein binding site of the promoter. We hypothesized that E1A might inhibit PEPCK gene transcription by binding to p300 and/or CBP. Surprisingly, we found that E1A consistently stimulated basal transcription from the PEPCK promoter in transfection assays in adenovirus (Ad)-infected HepG2 hepatoma cells or E1A-expressing, stably transfected 3T3 fibroblasts and nuclear run-on assays in Ad-infected H4IIE hepatoma cells. E1A also enhanced the stimulation of PEPCK gene transcription by Bt2cAMP. In transfection assays, wild type Ad5 expressing both 243R and 289R forms of E1A or a mutant virus expressing the 289R form alone stimulated transcription from the PEPCK promoter by approximately 5-fold 20 h postinfection. However, no stimulation was observed in cells infected with a virus expressing either the 243R protein alone or a 289R protein from which conserved region 3 (CR3) was mutated. Mutation or deletion of CR1 of E1A had no significant effect on transcription from the PEPCK promoter. Mutations within conserved region 2 (CR2) of E1A that inhibit the binding of E1A to the retinoblastoma gene product (pRb) further enhanced the stimulation of transcription from the PEPCK promoter by 2 3-fold compared with wild type E1A. These findings suggested that the normal function of pRb is to stimulate PEPCK gene transcription, and that this process is inhibited by the binding of E1A to pRb. This hypothesis was confirmed by overexpressing pRb in HepG2 cells, which stimulated transcription from the PEPCK promoter. Our findings indicate that Ad E1A regulates PEPCK gene transcription through a stimulatory mechanism involving CR3, and by attenuating a stimulatory effect of pRb through CR2.

Adenovirus E1A Proteins↗
PubMed1996

Inhibition of IFN-stimulated gene expression and IFN induction of cytolytic resistance to natural killer cell lysis correlate with E1A-p300 binding.

Treatment of target cells with IFN induces resistance to NK cell lysis. This process is blocked by expression of E1A gene products in adenovirus (Ad)-infected and Ad-transformed cells. We compared the ability of adenovirus serotype 5 (Ad5) E1A exon 1 mutants to inhibit the induction of cytolytic resistance by IFN and block IFN-stimulated gene expression with their capacity to bind the cellular proteins p105 (retinoblastoma gene product), p107, and p300. E1A mutants that did not express conserved region 3 (CR3; residues 138-184) or contained deletions in the nonconserved regions between residues 26-35 or 86-120, bound p105, p107, and p300 and were not impaired in their capacity to block IFN-stimulated gene expression or IFN's induction of cytolytic resistance. E1A mutants with deletions in CR2 (residues 121-138) could not bind p105 or p107, but blocked IFN-stimulated gene expression and IFN's induction of cytolytic resistance. In contrast, mutants in CR1 or the N-terminal nonconserved region (residues 2, 4-25, and 48-60), which define E1A's binding site for p300, were unable to block either IFN-stimulated gene expression or IFN's induction of cytolytic resistance. We conclude that E1A's capacity to block both IFN-stimulated gene expression and IFN's induction of cytolytic resistance appears to be transduced through a pathway that involves E1A-p300 binding. The capacity of E1A to block IFN's induction of cytolytic resistance is probably secondary to E1A's more general ability to inhibit IFN-stimulated gene expression.

Cytotoxicity, Immunologic↗
PubMed1995

E1A gene expression induces susceptibility to killing by NK cells following immortalization but not adenovirus infection of human cells.

Adenovirus (Ad) infection and E1A transfection were used to model changes in susceptibility to NK cell killing caused by transient vs stable E1A expression in human cells. Only stably transfected target cells exhibited cytolytic susceptibility, despite expression of equivalent levels of E1A proteins in Ad-infected targets. The inability of E1A gene products to induce cytolytic susceptibility during infection was not explained by an inhibitory effect of viral infection on otherwise susceptible target cells or by viral gene effects on class I MHC antigen expression on target cells. This differential effect of E1A expression on the cytolytic phenotypes of infected and stably transfected human cells suggests that human NK cells provide an effective immunologic barrier against the in vivo survival and neoplastic progression of E1A-immortalized cells that may emerge from the reservoir of persistently infected cells in the human host.

Adenovirus E1A Proteins↗
PubMed1995

Oncogenicity of human papillomavirus- or adenovirus-transformed cells correlates with resistance to lysis by natural killer cells.

The reasons for the dissimilar oncogenicities of human adenoviruses and human papillomaviruses (HPV) in humans are unknown but may relate to differences in the capacities of the E1A and E7 proteins to target cells for rejection by the host natural killer (NK) cell response. As one test of this hypothesis, we compared the abilities of E1A- and E7-expressing human fibroblastic or keratinocyte-derived human cells to be selectively killed by either unstimulated or interferon (IFN)-activated NK cells. Cells expressing the E1A oncoprotein were selectively killed by unstimulated NK cells, while the same parental cells but expressing the HPV type 16 (HPV-16) or HPV-18 E7 oncoprotein were resistant to NK cell lysis. The ability of IFN-activated NK cells to selectively kill virally transformed cells depends on IFN's ability to induce resistance to NK cell lysis in normal (i.e., non-viral oncogene-expressing) but not virally transformed cells. E1A blocked IFN's induction of cytolytic resistance, resulting in the selective lysis of adenovirus-transformed cells by IFN-activated NK cells. The extent of IFN-induced NK cell killing of E1A-expressing cells was proportional to the level of E1A expression and correlated with the ability of E1A to block IFN-stimulated gene expression in target cells. In contrast, E7 blocked neither IFN-stimulated gene expression nor IFN's induction of cytolytic resistance, thereby precluding the selective lysis of HPV-transformed cells by IFN-activated NK cells. In conclusion, E1A expression marks cells for destruction by the host NK cell response, whereas the E7 oncoprotein lacks this activity.

Adenovirus E1A Proteins↗
PubMed1994

Oral desensitization to rifampin and ethambutol in mycobacterial disease.

The incidence of disease caused by Mycobacterium tuberculosis (including drug-resistant strains) and M. avium complex (MAC) is increasing. Hypersensitivity reactions to antimycobacterial agents are relatively uncommon, but when they occur they may result in cessation of therapeutic medications. We report our experience with rapid oral desensitization to ethambutol and rifampin in a group of 10 patients with mycobacterial disease who had experienced cutaneous hypersensitivity reactions to these drugs. An adaptation of the rapid oral desensitization protocol for penicillin was used, with the dosing intervals increased to account for the different kinetics of these drugs. Adverse reactions were few and easily treated without necessitating cessation of therapy. We conclude that oral desensitization to rifampin and ethambutol by our protocol is safe and effective, allowing these patients to proceed with an optimal antimycobacterial regimen.

Administration, Oral↗
PubMed1993

Adenovirus E1A inhibits IFN-induced resistance to cytolysis by natural killer cells.

Infection of target cells with cytopathic viruses inhibits IFN induction of cytolytic resistance to NK cell-mediated cytolysis [IFN-mediated cytoprotection (IFN-MCP)]. It has been thought that the virally induced inhibition of IFN-MCP is secondary to the shutdown of cellular macromolecular synthesis that accompanies cytopathic virus infections. Group C, adenovirus serotype 5 (Ad5) infection inhibits both IFN-MCP and cellular protein synthesis. This study determined if the Ad5-induced inhibition of IFN-MCP was independent of adenovirus (Ad) infection and secondary only to the expression of the Ad early region 1A gene (E1A). To test this hypothesis, 4-h NK cytolysis assays were performed on IFN-gamma-treated human cells infected with an Ad5 E1A deletion mutant, dl343, or transfected with the Ad5 E1A gene. IFN-MCP was not inhibited by infection with dl343, despite the production of large amounts of both early (E1B, p55) and late (hexon) Ad proteins. In contrast to E1A-negative, parental cell lines, IFN-MCP was blocked in Ad5 E1A-transfected epithelial and fibroblastic cell lines. Genetic mapping studies within the E1A gene demonstrated that expression of only the first exon of E1A was sufficient to inhibit IFN-MCP. DNA sequence homology of E1A genes between different Ad groups (group A, Ad12; group C, Ad5) is limited almost entirely to three conserved regions located within the first exon of E1A. Because IFN-MCP was also blocked in Ad12 E1A-transfected cell lines, expression of one or more of the E1A-conserved regions may be necessary to inhibit IFN-MCP. In summary, the expression of E1A gene products inhibited IFN-MCP independently of virus infection. E1A's inhibition of IFN-MCP has the net effect of promoting the selective NK cell-mediated clearance of Ad-infected or Ad-transformed human cells.

Adenovirus E1A Proteins↗
PubMed1993

Activities of superoxide dismutases and NADPH oxidase in neutrophils obtained from asthmatic and normal donors.

PMN obtained from asthmatic subjects demonstrate a heightened respiratory burst with increased superoxide generation compared to normals. This enhanced superoxide anion generation could be secondary to increased activity of the respiratory burst NADPH oxidase or diminished metabolism of superoxide via superoxide dismutase (SOD). The two forms of SOD expressed in PMN, CuZnSOD expressed constitutively in the cytosol and inducible mitochondrial MnSOD, were investigated in asthmatics. Resting PMN from asthmatics (N = 9) contained significantly less MnSOD activity compared to controls (0.46 +/- 0.16 vs. 0.79 +/- 0.17 units/10(7) PMN, respectively; P = 0.0002). As several cytokines including interleukins (IL) -1, -4, and -6 as well as granulocyte macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor (TNF) enhance the PMN respiratory burst and are synthesized in the asthmatic lung, their effects on PMN MnSOD activity were assayed. In contrast to its effects on lymphocytes, both IL-1 and IL-6 significantly inhibited in a dose-dependent fashion the induction of MnSOD in PMN from normals (0.42 +/- 0.12 and 0.45 +/- 0.05 units/10(7) PMN, respectively, at 10 units/ml of each cytokine; P = 0.02 compared to resting cells) but failed to further modulate MnSOD production in asthmatic PMN. IL-4 and GM-CSF had no effect on MnSOD production, and TNF effects could not be studied because of its effects on cell viability. There were no differences in the activity of CuZnSOD (N = 9) or NADPH oxidase (N = 4) in the two groups. Inhibition of MnSOD activity in PMN secondary to cytokine exposure in the asthmatic lung could explain, at least in part, the increased generation of superoxide from PMN obtained from asthmatics. This would promote the presence and severity of inflammation in the asthmatic lung. These data further support a role for IL-1 and IL-6 in allergic inflammation.

Asthma↗
PubMed1993

Endogenous expression of E1A in human cells enhances the effect of adenovirus E3 on class I major histocompatibility complex antigen expression.

Group C human adenovirus (Ad) serotypes (e.g., Ad type 2 [Ad2] and Ad5) cause persistent infections in humans. One explanation for Ad persistence is an ineffective cytotoxic T-lymphocyte response due to diminished cell surface expression of class I major histocompatibility antigen (MHC Ag) on Ad-infected cells, an effect mediated by the Ad E3 19-kDa glycoprotein (E3 effect). However, we previously reported that, except for the Ad5 E1-transformed human cell line 293, a variety of human lymphoid, epithelial, and fibroblastic cells are resistant to the E3 effect during Ad5 infection (J. M. Routes and J. L. Cook, J. Immunol. 144:2763-2770, 1990). The present study tested the hypothesis that endogenous expression of E1A proteins in 293 cells sensitizes cells to this E3 effect, resulting in an enhanced downregulation of surface class I MHC Ag expression following Ad5 infection. Human epithelial and fibroblastic cells expressing E1A gene products for at least 72 h exhibited an enhanced E3 effect following Ad5 infection that was independent of baseline levels of surface class I MHC Ag expression and of E1A induction of E3 19-kDa glycoprotein expression. There was a direct correlation between the level of endogenous E1A expressed and the magnitude of the E3 effect. We postulate that the in vivo existence of cells stably expressing either E1A proteins or E1A-like activities in the microenvironment of Ad5 infection provides a reservoir of Ad-infected cells that is relatively protected from the virus-specific cytotoxic T-lymphocyte response, thereby favoring Ad persistence in humans.

Adenovirus E1A Proteins↗
PubMed1992

IFN increases class I MHC antigen expression on adenovirus-infected human cells without inducing resistance to natural killer cell killing.

It has been previously shown that unstimulated NK cells cannot preferentially lyse adenovirus serotypes 2 and 5-infected human cells. In this study, the ability of IFN to promote the selective NK cell-mediated lysis of adenovirus-infected human cells was determined. The relationship between target cell susceptibility to NK cell-mediated killing and class I Ag expression was also analyzed through the use of adenovirus serotype 2 and 5 mutants that do not make the adenovirus early region 3 19-kDa class I binding protein. IFN induced the selective lysis of adenovirus serotype 2 and 5-infected human cells by activating NK cells (IFN-alpha) and protecting uninfected, but not adenovirus-infected cells, from NK cell-mediated lysis (IFN-gamma). IFN-gamma increased the expression of class I Ag on the surface of cells infected with the adenovirus early region 3 deletion mutants, dl327 or dl801, to a level equal to or greater than that expressed on uninfected cells. Despite the increased expression of class I Ag, IFN-gamma could not protect these adenovirus-infected cells from NK cell-mediated lysis. Thus, dl327 or dl801 infection prevented IFN-gamma's induction of cytolytic resistance to NK cell-mediated killing but left IFN-gamma's induction of class I Ag intact. Surface class I Ag levels were substantially higher on IFN-gamma-treated, dl327-, and dl801-infected cells in comparison to cells infected with wild type adenovirus serotype 5. Again, higher target cell levels of class I Ag did not correlate with increased resistance to NK cell-mediated lysis because there was equivalent NK cell-mediated killing of IFN-gamma-treated adenovirus serotype 5-, dl327-, or dl801-infected cells. Thus, IFN-gamma only protects uninfected cells from NK cell-mediated killing, irrespective of target class I Ag levels, and thereby concentrates NK lytic activity on just adenovirus-infected cells. These data demonstrate that IFN-gamma's ability to protect target cells from NK cell-mediated cytolysis is unrelated to IFN-gamma's induction of surface class I MHC Ag.

Adenovirus Infections, Human↗