Biomedical subjects

D L Ewert

Publications and source records attributed to D L Ewert.

At least 37 records · Page 2Linked to original sources

PubMed1995

The ventriculo-arterial coupling ratio during transient Gz events.

It has been shown by Westerhof et al. that the ratio tau/T (where tau = Rp*SAC and Rp is total peripheral resistance, SAC is systemic arterial compliance, and T is heart period) is approximately a constant in all mammals under resting conditions such that diastolic pressure is sufficiently high to assure adequate coronary perfusion. The aim of this study is to determine if the ratio tau/T is constant under the transient condition of a rapid onset rate +Gz acceleration. We hypothesize that the ratio is held constant by the cardiovascular control system. Four male baboons were subjected to 10 second rapid onset +Gz profiles and aortic pressure and flow were recorded. Rp and C were calculated using a 2-element windkessel model and T was determined by the inverse of heart rate. All parameters were calculated on a beat-to-beat basis. It was found that the ratio increased very little at +2 Gz and increased dramatically at +3 and +5 Gz, disproving our hypothesis. This increase was due to large increases in peripheral resistance and changes in heart rate over the course of a Gz run. The increases are presumed to represent baroreceptor reflex response to the drop in aortic pressure experienced during +Gz stress. The ratio tau/T returned to its initial resting value shortly after the +Gz stressor was removed.

Animals↗

PubMed1994

A novel multilineage cell-surface antigen expressed on terminally differentiated chicken B cells in mucosal tissues.

A murine monoclonal antibody, 5M19, produced to a cell line transformed by an avian myeloblastosis virus, detects an antigen, designated chL5, that is expressed on multiple lineages of hematopoietic cells from noninfected chickens. Immunochemical analysis demonstrated the chL5 antigen to be a homodimer of two disulfide-bonded chains each having an apparent molecular weight of 128,000. The highest level of expression was found on myelomonocytic cells, including granulocytes, monocytes, and macrophages, as well as on activated T lymphocytes and plasma cells in mucosal tissues. A low level of chL5 expression is detectable on T lymphocytes in the thymus and peripheral tissues of juvenile chickens, and mitogen activation of T lymphocytes results in a twofold increase in antigen expression. By Day 15 of embryonic development chL5 is expressed by a small population (12%) of yolk sac cells and by the majority of mononuclear cells in the thymus, bone marrow, and spleen. Also, chL5+ cells populate the embryonic bursa of Fabricius before the appearance of IgM+ B lymphocytes. A novel characteristic of this antigen is its expression on about 50% of plasma cells in the mucosal tissues of the Harderian gland and cecal tonsils while present on only 3-11% of circulating B cells and less than 2% of splenic plasma cells. Immature B lymphocytes are negative for chL5 expression. Mitogen-induced activation and differentiation of peripheral B cells does not alter the percentage of chL5+ B cells. This antigen, therefore, is a useful marker for the analysis of terminal differentiation of B lymphocytes. The cell distribution of chL5 and the correlation of expression with cell aggregation in vitro suggests that the antigen may be involved in cell adhesion.

Animals↗

PubMed1994

Overexpression of C-terminally but not N-terminally truncated Myb induces fibrosarcomas: a novel nonhematopoietic target cell for the myb oncogene.

The myb oncogene encodes a DNA-binding transcriptional transactivator which can become a hematopoietic cell-transforming protein following the deletion of amino acid sequences from either its amino or carboxyl terminus. Although a number of hematopoietic tumors express terminally deleted variants of Myb, the involvement of truncated Myb in nonhematopoietic tumors has not been adequately investigated. To assess the full spectrum of Myb's oncogenic capability, a replication-competent retroviral vector (RCAMV) was used to express a full-length protein (C-Myb), an amino-terminally truncated protein (VCC- or delta N-Myb), a carboxyl-terminally truncated protein (T-Myb), or a doubly truncated protein (VCT-Myb) in vivo. These viruses were injected intravenously into 10-day chicken embryos, and the infected chicks were monitored for tumors. Approximately 4 to 8 weeks after hatching, the majority (30 of 39 [77%]) of animals infected with the T-Myb retrovirus (without 214 carboxyl-terminal residues) developed nodular muscle tumors which could be identified by both morphologic and immunohistochemical criteria as fibrosarcomas. Identically appearing tumors could also be found in the kidney of some T-Myb-infected animals. The T-Myb-induced fibrosarcomas expressed the appropriately sized T-Myb protein, contained an unaltered proviral T-myb gene, and showed clonal proviral integration sites. In comparison, no sarcomas were observed in any of the animals infected with the amino-terminally truncated (VCC- and delta N-Myb) or doubly truncated (VCT-Myb) viruses. A loss of carboxyl-terminal but not amino-terminal sequences can thus convert Myb into a potent in vivo transforming protein for nonhematopoietic mesenchymal cells. In comparison, a truncation of either or both ends of the protein can activate Myb into a hematopoietic cell-transforming protein.

Animals↗

PubMed1994

Beat-to-beat determination of peripheral resistance and arterial compliance during +Gz centrifugation.

This study focused on the problem of describing changes in total peripheral resistance (TPR) and systemic arterial compliance (SAC) under time-varying +Gz acceleration stress. Nonsteady-state measures of peripheral resistance can only be derived when arterial compliance is taken into account. We have developed a successful analytical model to track simultaneous changes in peripheral resistance and systemic arterial compliance during non-stationary periods of increased gravitational load on a beat-to-beat basis. Using a 2-element windkessel model, aortic flow into an input node was defined as equal to the sum of a capacitative (Cao) and a resistive (Rarterial) flow leaving the node such that: Iao = Caod(Pao - Ppleural)/dt + (Pao - Pra)/Rarterial We made the assumption that Cao and Rarterial were constant over a cardiac cycle, and divided the pressure and flow signals for each beat of a record into two different intervals, integrating this equation over each, giving two equations in two unknowns. Cao and Rarterial were then obtained from the matrix solutions. To test the model, we used recordings from chronically instrumented baboons subjected to a 10 s rapid onset +Gz (head-to-foot) stress. Beat-to-beat calculations of peripheral resistance and systemic arterial compliance from our model were compared to values obtained from a previously reported 3-element wind-kessel model.

Animals↗

PubMed1994

Bcl-2 expressed using a retroviral vector is localized primarily in the nuclear membrane and the endoplasmic reticulum of chicken embryo fibroblasts.

A complementary DNA for human bcl-2 was cloned into the replication competent avian retrovirus vector RCASBP, and the resulting virus was used to express human Bcl-2 protein at high levels in chicken embryo fibroblasts. The expression of Bcl-2 did not transform or significantly alter the longevity of the chicken embryo fibroblasts in the presence of normal amounts of serum. However, the expression of Bcl-2 blocked c-Myc-induced apoptosis in these cells. Fractionation of the infected chicken embryo fibroblasts indicated that the protein was distributed equally between nuclear and high density cytoplasmic membranes. Immunofluorescence analysis by confocal microscopy and immunoelectron microscopy showed that the Bcl-2 protein was primarily associated with the nuclear membrane and with the endoplasmic reticulum. Reduced amounts of the protein were associated with other membranes in the cytoplasm. These data show that, in this system, the Bcl-2 protein associates with the nuclear membrane and intracytoplasmic membranes but is not preferentially associated with mitochondria.

Animals↗

PubMed1993

Expression and role of c-myc in chondrocytes undergoing endochondral ossification.

To analyze the relationship between c-myc gene expression and chondrocyte proliferation and maturation during endochondral ossification, Day 18-19 chick embryo sterna were pulse-labeled with [3H]thymidine, and serial sections were processed for autoradiography and in situ hybridization. Proliferating chondrocytes, located in four distinct areas of the developing sternum, all contained high levels of c-myc transcripts, whereas postmitotic chondrocytes (such as hypertrophic chondrocytes) contained undetectable amounts. These findings were confirmed by Northern blot analysis and by the observation that antisense c-myc oligomer treatment inhibited proliferation in cultured chondrocytes. Constitutive overexpression of c-myc by retroviral vectors in immature chondrocyte cultures (c-myc cultures) maintained the cells in a proliferative state and blocked their maturation into hypertrophic chondrocytes. The lack of maturation in the c-myc cultures was corroborated by analysis of type X collagen gene regulation. Control immature cultures contained strong repressor activity for the type X collagen gene promoter, as revealed by transfection assays; repressor activity was lost upon maturation and activation of type X collagen synthesis. In the c-myc cultures, however, repressor activity persisted. Thus, c-myc participates in the normal changes in proliferation accompanying chondrocyte maturation in vivo and in culture. The decreases in c-myc expression and cell proliferation appear to be required for completion of maturation.

Animals↗

PubMed1993

Latency and reactivation of Marek's disease virus in B lymphocytes transformed by avian leukosis virus.

The physical and biological state of the Marek's disease virus (MDV) genome in avian leukosis virus (ALV)-transformed cells is characterized using cell lines established from ALV tumours co-infected with the SB-1 strain of MDV. The MDV genome within the ALV-transformed cells was found to be methylated at 5' CpG 3' dinucleotides. Less than 2% of the tumour cells expressed MDV antigen and only one virus plaque that was characteristic of an MDV infection was noted when tumour cells were cocultured with fibroblasts permissive for a productive MDV infection. However, when methylation of the MDV genome was prevented by culturing the tumour cell lines in the presence of 5-azacytidine, both MDV antigen expression and viral replication increased. Based on these results, it appears that MDV resides within the ALV-transformed cells in a latent state and that MDV latency might be influenced, to some extent, by methylation of the MDV genome.

Alpharetrovirus↗

PubMed1992

Isolation and characterization of the chicken bcl-2 gene: expression in a variety of tissues including lymphoid and neuronal organs in adult and embryo.

The expression of human bcl-2 gene is de-regulated by t(14;18) translocation in most of follicular lymphoma. Recent studies indicated that the bcl-2 gene product has an ability to block apoptosis of hematopoietic cells. To facilitate the analysis of the role of this gene in normal development using an animal model, we have isolated and partially characterized the chicken homologue of human bcl-2 gene. The analysis of nucleotide sequence showed that the organization of the chicken bcl-2 gene is very similar to that of human bcl-2 gene. The primary transcript is spliced to encode a 25,687 dalton (233 a.a.) protein. The chicken Bcl-2 protein has two regions highly homologous to human Bcl-2 protein surrounding a totally non-homologous region. The expression of the chicken bcl-2 gene was analyzed in various chicken tissues. In the adult chicken, bcl-2 transcripts were detected in thymus, spleen, kidney, heart, ovary and brain, with the highest levels being detected in the thymus. However, the bursa of Fabricius, which is the site of early B cell development, expressed much less amounts of bcl-2 RNA. On the other hand, in embryo, the gene is extensively expressed in the bursa, as well as in muscle and the above tissues. Our findings indicate that a homologue of the human bcl-2 gene does exist in the chicken and that its expression is developmentally regulated in some tissues.

Amino Acid Sequence↗

PubMed1992

Major histocompatibility (B) complex control of the growth pattern of v-src DNA-induced primary tumors.

Observations that the major histocompatibility (B) complex is a determinant of the growth pattern of Rous sarcoma virus (RSV)-induced tumors raised the question as to whether control is exerted at the level of a v-src-determined, i.e., transformation-specific, function. To investigate this point, the tumor size scores and tumor profile indices of v-src-induced tumors were compared in two lines of chickens congenic for B complex genotypes. The finding that the growth patterns of tumors, induced by v-src DNA inoculation at 6 weeks posthatch, differ in these two lines establishes that the B complex exerts control over tumor growth at the level of a v-src-determined function. The potential importance of this control, in terms of the naturally occurring case of an avian sarcoma virus infection, is suggested by the observation that the patterns of tumor growth in a given congenic line are similar whether the tumors are induced by v-src DNA or by RSV.

Animals↗

PubMed1992

Persistence of Marek's disease virus in a subpopulation of B cells that is transformed by avian leukosis virus, but not in normal bursal B cells.

Previous studies have described an augmentation of avian leukosis virus (ALV)-induced lymphoid leukosis in chickens that were coinfected with a serotype 2 Marek's disease virus (MDV) strain, SB-1. As a first step toward understanding the mechanism of this augmentation, we have analyzed the tropism of the MDV for the ALV-transformed B cell. After hatching, chickens were coinfected with ALV and a nonpathogenic strain of MDV, SB-1. Seventy primary and metastatic ALV-induced lymphomas that developed in chickens between 14 and 20 weeks of age were found, with only one exception, to carry SB-1 DNA. The MDV genome was maintained in cell lines derived from the tumors. However, MDV DNA could not be detected in nontransformed bursal B cells from chickens carrying ALV lymphomas. Moreover, during and after the lytic phase of MDV infection, SB-1 DNA was near or below the level of detection in bursal cells, suggesting that MDV most likely infects only a small subpopulation of bursal cells. By contrast, ALV-transformed B cells from MDV-free chickens could be persistently infected with MDV in vitro. These findings indicate that ALV lymphoma cells, unlike nontransformed bursal B cells, are susceptible to persistent MDV infection and can serve as a reservoir of MDV that can potentially influence the physiology of the transformed cell.

Animals↗

PubMed1992

Transformation of chicken myelomonocytic cells by a retrovirus expressing the v-myb oncogene from the long terminal repeats of avian myeloblastosis virus but not Rous sarcoma virus.

To test the effect of long terminal repeat (LTR) regulatory sequences on the transforming capability of the v-myb oncogene from avian myeloblastosis virus (AMV), we have constructed replication-competent avian retroviral vectors with nearly identical structural genes that express v-myb from either AMV or Rous sarcoma virus (RSV) LTRs. After transfection into chicken embryo fibroblasts, virus-containing cell supernatants were used to infect chicken myelomonocytic target cells from preparations of 16-day-old embryonic spleen cells. Both wild-type AMV and the virus expressing v-myb from AMV LTRs (RCAMV-v-myb) were able to transform the splenocyte cultures into a population of immature myelomonocytic cells. The transformed cells expressed the p48v-Myb oncoprotein and formed compact foci when grown in soft agar. In contrast, the virus expressing v-myb from RSV LTRs (RCAS-v-myb) was repeatedly unable to transform the same splenocyte cells, despite being able to infect fibroblasts with high efficiency. This difference in the transforming activities of v-myb-expressing viruses with different LTRs most likely results from the presence of a factor (or factors) within the appropriate myelomonocytic target cell that promotes specific expression from the AMV but not from the RSV LTR.

Animals↗

PubMed1992

In vivo relation of intramyocardial blood volume to myocardial perfusion. Evidence supporting microvascular site for autoregulation.

BACKGROUND: The goal of this study was to explore the role of several factors that affect intramyocardial blood volume by using minimally invasive computed tomography. Anesthetized dogs were scanned with the dynamic spatial reconstructor, a high-speed tomographic scanner, during injection of a bolus of iohexol into the aortic root. METHODS AND RESULTS: In control dogs, it is indicated that the fraction of myocardium that is blood (FMB, %) relates to myocardial perfusion (F, milliliters per gram per minute) in that region as FMB congruent to a.F1/2, where a = 9.5 +/- 1.2% (milliliters times minute per gram)1/2 (mean +/- SD) in the subendocardium and a = 9.6 +/- 1.1% in the subepicardium. In another group of dogs, for the myocardium perfused by a stenosed epicardial artery, a increased to approximately 10 for a 25-43% stenosis (or pressure gradient of 9 mm Hg across narrowing) and to greater than 11 for a 50-55% stenosis (or pressure gradient of 40 mm Hg across narrowing). In these dogs, flow was not impaired under control hemodynamic conditions, but the usual increase of flow (i.e., flow reserve) observed under maximum vasodilation conditions was impaired. In another group of dogs, progressive embolization (using 15-microns-diameter microspheres) of the left ventricular myocardial microcirculation caused the value of a to remain at approximately 9.5 with embolization up to 50% of the fatal dose of microspheres, but it then decreased progressively with embolization to 4.6 at the fatal dose. CONCLUSIONS: We conclude that the FMB/F relation reflects hemodynamic conductance at the microvascular level.

Animals↗

PubMed1992

Retroviruses that express different ras mutants cause different types of tumors in chickens.

We have used replication-competent retroviral vectors to express avian and murine ras genes in cultured chick embryo fibroblasts (CEF) and in chickens. Since the viral vectors are identical, it is possible to compare the oncogenic potential of the ras genes directly. The normal (12 gly) form of chicken c-Ha-ras is not oncogenic in vivo, nor does high-level expression transform CEF. Expression of murine v-ras or modified forms of chicken c-Ha-ras with either lysine or glutamine at position 12 transforms CEF and causes tumors in birds. However, the oncogenic potential of the transforming ras genes is different; the viruses that express the genes with lysine and glutamine at position 12 cause a distinct spectrum of tumors.

Animals↗

PubMed1991

Induction of a diffuse mesothelioma in chickens by intraperitoneal inoculation of v-src DNA.

The presence of peritoneum-based, cytokeratin-positive tumor cells, as detected under conditions of intravenous inoculation of chickens with a v-src-positive DNA fragment, raised the possibility that such DNA may be inductive for diffuse mesothelioma. To investigate this possibility, chickens were inoculated intraperitoneally with v-src DNA, and the resultant peritoneum-based tumor tissue was subjected to histopathological analysis. On the basis of (i) morphology, (ii) immunohistochemistry/histochemistry, and (iii) ultrastructure, we concluded that this tissue represents a diffuse mesothelioma.

Animals↗

PubMed1990

Light chain gene conversion continues at high rate in an ALV-induced cell line.

We have analyzed immunoglobulin light chain sequences from avian leukosis virus (ALV) induced bursal and metastatic tumors and from cell lines derived from these tumors. Sequence data presented demonstrate that ALV-induced tumors and one cell line (DT40) derived therefrom continue to diversify their light chain genes outside of the bursal environment. Diversification within these tumor cells seems to occur by gene conversion events comparable with those observed in bursal B cells. Sequence analysis of spontaneously arising surface immunoglobulin negative subclones of the DT40 cell line revealed frameshifts within the rearranged light chain genes which most likely resulted from non-functional recombination events. Superimposed gene conversion events can repair these frameshifts leading to re-expression of surface immunoglobulin.

Animals↗

PubMed1990

Wing web or intravenous inoculation of chickens with v-src DNA induces visceral sarcomas.

We have previously found that sarcomas localized to visceral organs frequently arise following wing web inoculation of Rous sarcoma virus. This observation prompted an investigation of whether visceral sarcomas are also inducible by wing web inoculation of a subgenomic viral DNA fragment (v-src DNA) that includes v-src but lacks genes encoding viral structural protein. For this analysis, line SC chickens were inoculated with v-src DNA at 1-2 days posthatch and monitored for 9 weeks. Ninety percent of the chickens developed wing web (primary) tumors at the site of inoculation and, of these, about 30% exhibited visceral tumors. All tumors were histologically identifiable as sarcomas, and both the primary and visceral sarcoma cells were specifically reactive with two monoclonal antibodies elicited to different peptide fragments of pp60v-src. In a separate set of experiments, visceral sarcomas were also observed in about half of the line SC chickens that had been inoculated intravenously with v-src DNA. These results indicate that exogenous progeny virus production is not required for v-src-induced, visceral sarcoma formation. In addition, they demonstrate that intravenous inoculation of v-src DNA is a means to achieve rapid and widespread, disseminated sarcoma growth.

Animals↗

PubMed1990

Markers of B lymphocyte differentiation in the chicken.

A study was made of the ontogeny and tissue distribution of seven antigen systems associated with B lymphocyte development. A panel of seven monoclonal antibodies (MAbs) directed against avian bursal and peripheral B lymphocytes was developed. The spectra of cellular reactivity and size of respective antigens indicate that these MAbs appear to react with determinants of chicken B lymphocytes that had not been previously described. Immunofluorescence analysis of cell surface antigens using this panel indicated that dynamic changes in antigen expression are associated with early ontogeny and maturation of the B cell lineage. The yolk sac contained subpopulations of hematopoietic cells reactive with three MAbs (CB10, CLA3 and Hy86b5) which possibly mark the pre-bursal stem cell population. At or near the time of surface IgM expression in the embryonic bursa, B cells expressed antigens detected by the CB7, CB8, CB9 and CB11. In the juvenile chicken, CB7 reacted with immature bursal lymphocytes, while CB8, CB9 and CB10 reacted with bursal lymphocytes and a subpopulation of peripheral B lymphocytes. The antigens defined by Hy86b5 and CB11 were expressed on mature B lymphocytes but on only a subpopulation of bursal lymphocytes. The MAb CLA3 detected an antigen expressed on all leukocytes but not erythroid cells. Expression of some of these antigens by cells of the myelomonocytic lineages suggests a possible functional or ontogenetic relationship between the B lymphocyte and myeloid lineages.

Animals↗

PubMed1990

Reticuloendotheliosis virus REV-T(REV-A)-induced neoplasia: development of tumors within the T-lymphoid and myeloid lineages.

Infection of 1-day-old chicks with reticuloendotheliosis virus strain T induces a neoplastic disease that kills the chicks 7 to 14 days postinfection. In association with reticuloendotheliosis-associated virus (REV-A), reticuloendotheliosis virus T (REV-T) induces tumors that are predominantly immunoglobulin M (IgM) negative. We examined a variety of REV-T(REV-A)-induced tumors and tumor-derived cell lines and concluded that the principal IgM-negative tumors that develop in REV-T(REV-A)-infected chicks are neither pre-B or pre-B-pre-T but rather mature T lymphoid and myeloid. Without exception, the immunoglobulin heavy- and light-chain loci were in germ line configuration. Furthermore, the cell lines expressed neither sterile transcripts of the heavy- or light-chain immunoglobulin genes nor elevated levels of c-myb, two characteristics associated with murine pre-B lymphomas. Cell lines were also examined by using monoclonal antibodies for expression of a variety of cell surface markers expressed on B lymphocytes and/or T lymphocytes and/or myeloid cells. These reagents defined two types of IgM-negative tumor cell lines, one CIa+ CT-3+ (T lymphoid) and the other CIa+ CT-3-. By using the same approaches, tumor development was examined following REV-T(REV-A) infection at 1 and 3 weeks post-hatching of cyclophosphamide-treated chicks shown to be devoid of B-lymphoid cells. Again, the tumors that developed were either CIa+ CT-3+ (T lymphoid) or CIa+ CT-3-. Furthermore, the frequency and rate with which IgM-negative tumors developed in cyclophosphamide-treated chicks were not different from those observed in normal chicks. In 3-week-old cyclophosphamide-treated chicks, the presence of CIa+ CT-3- tumors bearing hematopoietic lineage markers, such as CLA-3 and 5M19, are most likely to have been derived from cells within the myeloid lineage.

Animals↗