Search PubMed⌕ Search

Biomedical subjects

G R Johnson

Publications and source records attributed to G R Johnson.

At least 19 recordsLinked to original sources
PubMed1995

Murine hematopoietic stem and progenitor cells: I. Enrichment and biologic characterization.

Murine bone marrow cells were fractionated by fluorescence-activated cell sorting into Rh123lo Lin- c-kit+ Ly6A+, Rh123hi Lin-c-kit+ Ly6A+, and Lin- c-kit+ Ly6A- populations within which most, if not all, of the hematopoietic activities of the marrow resided. The Rh123lo Lin- c-kit+Ly6A+ cells, which consist exclusively of small- or medium-sized lymphocyte-like cells, are highly enriched for long-term hematopoietic in vivo repopulating cells. The enrichment factor for these cells from the marrow was estimated as 2,000-fold. The Rh123hi Lin- c-kit+ Ly6A+ cells, although also highly enriched for day-12 spleen colony-forming units, were relatively depleted of long-term in vivo repopulation capacity. Most, if not all Lin- c-kit+ Ly6A- cells were Rb123hi. In contrast to both Rh123lo and Rh123hi Lin- c-kit+ Ly6A+ stem cell populations, the Lin- c-kit+ Ly6A- cells can be stimulated to proliferate in vitro in the presence of single cytokines, which is a characteristic of committed progenitor cells. No marked synergistic interactions between individual cytokines were observed with this cell population. Both Rh123hi Lin- c-kit+ Ly6A+ mature stem cell and Lin- c-kit+ Ly6A- progenitor cell populations displayed in vivo repopulation kinetics resembling those of the putative short-term hematopoietic repopulating cells.

Animals↗
PubMed1995

mRNA phenotyping of the major ligands and receptors of the EGF supergene family in human ovarian epithelial cells.

mRNA amplification phenotyping (MAPPing) was used to determine the level of mRNA expression of the major EGF-related ligands (EGF, TGF-alpha, and Amphiregulin) and receptors (EGF-receptor and erbB-2) of the EGF supergene family in three ovarian carcinoma lines (OVCA 429 and 433, and NIH:OVCAR-8) under serum-supplemented and reduced serum (minimal medium with 2% fetuin) growth conditions. mRNA levels of TGF-alpha, EGF-R, and erbB-2 were particularly high, and increased approximately 2-3 orders of magnitude when grown in serum, consistent with an autocrine involvement of these genes in ovarian epithelial growth in vitro. Moreover, even when grown without serum, OVCA 429 and NIH:OVCAR-8 expressed elevated levels of mRNA for erbB-2.

Amphiregulin↗
PubMed1995

Localization of amphiregulin in the human placenta and decidua throughout gestation: role in trophoblast growth.

Amphiregulin (AR) is a growth regulatory glycoprotein with significant amino acid homology to members of the epidermal growth factor (EGF) family. Its effects are mediated via the EGF receptor tyrosine kinase or through specific nuclear targeting sequences. In this study, the localization of immunoreactive AR was examined in paraformaldehyde-glutaraldehyde fixed, paraffin-embedded human placentae and decidua obtained at various stages from 11 weeks to term pregnancy using the avidin-biotin complex/peroxidase method. In addition, the effects of AR on trophoblast proliferation were evaluated from 3HTdR uptake by first trimester human trophoblast cells. Results revealed immunolocalization of AR to the nucleus as well as the cytoplasm of the syncytiotrophoblast cell layer of chorionic villi until approximately week 18 of gestation after which no immunostaining was detected. Villous and extravillous cytotrophoblast cells as well as decidual tissue were negative for AR at all gestational ages examined. In the presence of exogenous AR, there was a dose-dependent increase in proliferation of the trophoblast at AR concentrations ranging from 1-100 ng/ml. These results suggest that AR may be an important paracrine or juxtacrine growth stimulatory molecule for cytotrophoblast cells in situ early in gestation.

Amphiregulin↗
PubMed1995

Variability in measurement of abdominal aortic aneurysms. Abdominal Aortic Aneurysm Detection and Management Veterans Administration Cooperative Study Group.

PURPOSE: The purpose of this study was to report interobserver and intraobserver variability of computed tomography (CT) measurements of abdominal aortic aneurysm (AAA) diameter and agreement between CT and ultrasonography observed in the course of a large, multicenter, randomized trial on the management of small AAAs. METHODS: CT measurements of AAA diameter from participating centers were compared with measurements made from the same scan by a central laboratory. Blinded central remeasurement of a randomly selected subset of these CT scans was used to assess intraobserver variability. Agreement between AAA measurements by CT and ultrasonography done within 30 days of each other was also assessed. RESULTS: For interobserver pairs of local and central CT measurements of AAA diameter (n = 806), the difference was 0.2 cm or less in 65% of pairs, but 17% differed by at least 0.5 cm. For intraobserver pairs of central CT remeasurements (n = 70), 90% differed by 0.2 cm or less, 70% were within 0.1 cm, and only one differed by 0.5 cm. Of 258 ultrasound-measured and central CT pairs, the difference was 0.2 cm or less in 44% and at least 0.5 cm in 33%. Ultrasound measurements were smaller than central CT measurements by an average of 0.27 cm (p < 0.0001). Local CT and ultrasound measurements showed a marked preference for recording by half centimeter. CONCLUSIONS: A high degree of precision is possible in CT measurement of AAA diameter, but this precision may not be obtained in practice because of differences in measurement techniques. Differences between imaging modalities increase variability further. Variations in AAA measurement of 0.5 cm or more are not uncommon, and this should be taken into account in management decisions. Efforts to reduce variation in measurement are warranted and might include (1) seeking agreement between surgeons and radiologists on a precise definition of AAA diameter, (2) limiting the number of radiologists who measure AAAs, and (3) use of calipers and magnifying glass for CT measurements.

Aged↗
PubMed1995

Nucleotide sequence analysis of genes encoding a toluene/benzene-2-monooxygenase from Pseudomonas sp. strain JS150.

It was previously shown by others that Pseudomonas sp. strain JS150 metabolizes benzene and alkyl- and chloro-substituted benzenes by using dioxygenase-initiated pathways coupled with multiple downstream metabolic pathways to accommodate catechol metabolism. By cloning genes encoding benzene-degradative enzymes, we found that strain JS150 also carries genes for a toluene/benzene-2-monooxygenase. The gene cluster encoding a 2-monooxygenase and its cognate regulator was cloned from a plasmid carried by strain JS150. Oxygen (18O2) incorporation experiments using Pseudomonas aeruginosa strains that carried the cloned genes confirmed that toluene hydroxylation was catalyzed through an authentic monooxygenase reaction to yield ortho-cresol. Regions encoding the toluene-2-monooxygenase and regulatory gene product were localized in two regions of the cloned fragment. The nucleotide sequence of the toluene/benzene-2-monooxygenase locus was determined. Analysis of this sequence revealed six open reading frames that were then designated tbmA, tbmB, tbmC, tbmD, tbmE, and tbmF. The deduced amino acid sequences for these genes showed the presence of motifs similar to well-conserved functional domains of multicomponent oxygenases. This analysis allowed the tentative identification of two terminal oxygenase subunits (TbmB and TbmD) and an electron transport protein (TbmF) for the monooxygenase enzyme. In addition to these gene products, all the tbm polypeptides shared significant homology with protein components from other bacterial multicomponent monooxygenases. Overall, the tbm gene products shared greater similarity with polypeptides from the phenol hydroxylases of Pseudomonas putida CF600, P35X, and BH than with those from the toluene monooxygenases of Pseudomonas mendocina KR1 and Burkholderia (Pseudomonas) pickettii PKO1. The relationship found between the phenol hydroxylases and a toluene-2-monooxygenase, characterized in this study for the first time at the nucleotide sequence level, suggested that DNA probes used for surveys of environmental populations should be carefully selected to reflect DNA sequences corresponding to the metabolic pathway of interest.

Amino Acid Sequence↗
PubMed1995

Estrogen and phorbol esters regulate amphiregulin expression by two separate mechanisms in human breast cancer cell lines.

The actions of 17 beta-estradiol (E2) and protein kinase C (PKC) appear to converge in the regulation of expression of certain growth modulatory genes, such as the growth factor amphiregulin (AR). AR is known to modulate cell growth by binding to the epidermal growth factor receptor. In the current report we established the mechanisms of the PKC-activating phorbol ester tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA) and the steroid hormone E2 on the induction of AR expression in human breast carcinoma cell lines. TPA (100 nM) and E2 (1 nM) induce AR messenger RNA (mRNA) expression by 6- to 8-fold and 3- to 6-fold, respectively, in a time- and dose-dependent manner. In addition, immunoreactive AR protein is induced by both TPA and E2 by 6- to 8-fold and 2- to 4-fold, respectively. The PKC-modulating drugs, bryostatin and H-7, and antiestrogens (ICI 164,384 and 4-hydroxytamoxifen) interfere with AR induction by TPA and estrogen, respectively. The effects of TPA and E2 on the induction of AR mRNA were both closely associated with enhanced transcription of the AR gene. However, TPA had an additional effect at the posttranscriptional level by stabilizing the AR mRNA. The protein synthesis inhibitor, cycloheximide, prevented AR induction by TPA, suggesting that a component of the TPA induction of AR is indirect and dependent upon protein synthesis. Conversely, the E2 induction of AR transcription was found to be a direct response, independent of protein synthesis. The results presented herein thus demonstrate that TPA and E2 are able to stimulate AR gene transcription by two separate mechanisms.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1995

Syndrome of flank pain and acute renal failure after binge drinking and nonsteroidal anti-inflammatory drug ingestion.

The binge drinking of alcohol combined with the ingestion of a nonsteroidal anti-inflammatory drug (NSAID) is a recently described cause of reversible acute renal failure. The pathogenetic mechanisms leading to acute tubular necrosis in this setting include the initial compromise in renal perfusion due to alcohol-induced extracellular volume contraction and the superimposed renal hemodynamic alterations induced by the NSAID that interfere with the renal autoregulation. Although alcohol may also cause rhabdomyolysis leading to acute tubular necrosis, this is usually not apparent in these cases. Previously, only three such cases have been reported but the incidence is likely to be higher in view of the prevalence of alcohol and NSAID use. Herein is presented another patient in whom the features of flank pain and acute renal failure in association with binge drinking and NSAID ingestion constitute a characteristic syndrome.

Abdominal Pain↗
PubMed1995

Role of talectomy in the treatment of rigid talipes equinovarus deformities.

The authors provide a review of the principles of talectomy as a surgical treatment option for severe, rigid, and resistant clubfoot deformities. Such deformities are associated with a high recurrence rate and frequently involve a large number of surgical interventions to attain a satisfactory result. Talectomy has been successfully used as both a primary and salvage procedure to treat such deformities, often eliminating the need for subsequent surgeries. A discussion of the operative technique, indications, advantages, and common complications associated with the procedure is presented. In addition, a case report of talectomy used to treat a severe, bilateral clubfoot deformity in a 4-year-old boy is included.

Child, Preschool↗
PubMed1995

Lineage-restricted recruitment of immature hematopoietic progenitor cells in response to Epo after normal hematopoietic cell transfection with EpoR.

The receptor for erythropoietin (EpoR) is normally restricted in its expression to the relatively mature cells of the erythroid and megakaryocytic lineages. Using retrovirus-mediated gene transfer, the wild-type EpoR and a constitutively activated mutant of the EpoR, EpoRR129C, were expressed in primary hematopoietic cells. Retroviral infection of day-12 murine fetal liver, followed by stimulation with Epo as a single stimulus, generated day-8 erythroid colonies resembling colonies derived from burst-forming units-erythroid (BFU-E). Similarly, murine post-5 fluorouracil (5-FU) bone marrow cells or fetal liver cells, induced to express EpoR and stimulated by Epo, displayed a significant enhancement of megakaryocyte colony formation, particularly of the BFU-megakaryocyte (BFU-Mk) colony type. Cultures of bone marrow cells transduced with the EpoR retrovirus and stimulated by Epo contained macrophage colonies but very few granulocyte colonies. Experiments to culture single clones demonstrated direct action of Epo on megakaryocyte and macrophage clones but failed to demonstrate a direct action on granulocyte precursors. A similar pattern of lineage-restricted effects was demonstrated in unstimulated cultures of cells infected with the EpoRR129C retrovirus. In summary, we have demonstrated Epo-induced recruitment of immature erythroid and megakaryocyte precursors induced to express the EpoR. Furthermore, we have also demonstrated lineage-restricted cell proliferation in response to Epo by normal myeloid hematopoietic cells transduced with the EpoR.

Animals↗
PubMed1995

Overexpression of C-FMS in the myeloid cell line FDC-P1 induces transformation that dissociates M-CSF-induced proliferation and differentiation.

Hemopoietic growth factors have been implicated in the pathogenesis of some myeloid leukemias when the cells acquire the autocrine capacity to produce a relevant growth factor. We report transformation of the immortalized cell line FDC-P1 by overexpression of murine c-fms, the receptor for macrophage colony-stimulating factor (M-CSF). Three types of cell lines were obtained following expression of c-fms in FDC-P1 cells. The first type of cell line (FD-c-fms) was dependent on stimulation by M-CSF to induce cell proliferation and also underwent limited monocytic/macrophage differentiation in response to M-CSF. The second type of cell line (FD-c-fmsi) was able to proliferate in serum-containing cultures without stimulation by exogenous hemopoietic regulators. Stimulation of the FD-c-fmsi cell lines with M-CSF suppressed proliferation compared to cultures of unstimulated cells and induced monocytic/macrophage differentiation. The third type of cell line (FD-c-fms) was autonomous, failed to differentiate in response to M-CSF and secreted M-CSF into the culture medium, suggesting an autocrine mechanism of transformation. All three types of cell lines demonstrated varying degrees of suppression of cell proliferation when stimulated by the combination of M-CSF and GM-CSF. The data indicate that transformation of c-fms expressing cells results in a variety of phenotypes with dissociation of M-CSF induced proliferation and differentiation in different cell lines.

Cell Differentiation↗
PubMed1995

Purified murine long-term in vivo hematopoietic repopulating cells are not prothymocytes.

Analysis of kinetics of thymic repopulation by Rh123low, Lin-, Ly6A/E+, c-kit+ (Rh123low) cells, highly enriched for long-term in vivo hematopoietic repopulating cells, reveals that this population is deficient in thymic repopulation at week 3 after intravenous transplantation when compared to normal bone marrow cells. This suggests that the marrow prothymocytes have been depleted from this population, and analysis of thymic repopulation at week 3 can therefore be used to differentiate prothymocytes and their precursors. Using this short-term assay, the Rh123high, Lin-, Ly6A/E+, c-kit+ (Rh123high) population has been found to be relatively more efficient at early thymic repopulation, suggesting that this population contains the prothymocytes. In addition, the differentiation potential and reconstitution behavior of the Rh123high population observed after intravenous and intrathymic transfer strongly indicates that this population is at the transitional stage between the marrow primitive pluripotential and thymic more mature lymphoid restricted stem cells. We propose that the thymic repopulating ability of the Rh123low population is through generation of the more mature Rh123high progeny, presumably in the marrow.

Animals↗
PubMed1994

Heparan sulfate is essential to amphiregulin-induced mitogenic signaling by the epidermal growth factor receptor.

Human amphiregulin (AR) is a heparin-binding growth factor which functions by binding to and activating the epidermal growth factor (EGF) receptor tyrosine kinase. AR contains an EGF-like domain (residues 44-84) and a Lys/Arg-rich NH2-terminal extension (residues 1-43). Synthetic peptides corresponding to residues 8-26, 26-44, and 68-84 of AR were tested for their ability to compete for the binding of AR to immobilized heparin. AR8-26 and AR68-84 had no significant effect on the binding of AR to heparin, whereas AR26-44 bound to heparin and blocked the binding of AR to heparin. Both soluble heparin and heparan sulfate inhibited AR-induced mitogenesis in MCF-10A human mammary epithelial cells with an IC50 of 5 and 2 micrograms/ml, respectively, whereas soluble chondroitin sulfate had only a slight inhibitory effect. When MCF-10A cells were grown in the presence of chlorate, an inhibitor of sulfation, or exposed to the glycosaminoglycan-degrading enzymes heparitinase or heparinase, the ability of AR to evoke mitogenesis in these cells was lost. Chlorate, heparitinase, or heparinase treatment inhibited AR-induced autophosphorylation of tyrosine residues in the EGF receptor. None of these treatments had any significant effect on EGF-triggered mitogenic signaling by the EGF receptor. These results indicate that extracellular heparan sulfate glycosaminoglycan is essential to AR-induced mitogenic signaling by the EGF receptor tyrosine kinase.

Amphiregulin↗
PubMed1994

Induction and expression of amphiregulin in human pancreatic cancer.

The epidermal growth factor receptor is activated by a family of polypeptides that includes the growth factor amphiregullin (AR). Using Northern blot analysis and the polymerase chain reaction, we now report that AR mRNA is expressed in human pancreatic cancer cell lines, and that this expression is enhanced in several of these cell lines by tetradecanoyl phorbol acetate and transforming growth factor alpha. AR was also expressed in normal and malignant pancreatic tissues. However, in the normal pancreas, AR immunostaining was most evident in the nuclei of ductal cells. In contrast, in many carcinomas, AR was also present in the cytoplasm of the ductal-like cancer cells. Cytoplasmic localization of AR was associated with a more advanced clinical stage. These findings suggest that AR may contribute to aberrant activation of the epidermal growth factor receptor in human pancreatic cancer, and may enhance disease progression.

Amino Acid Sequence↗
PubMed1994

Stem cell factor enhances the survival but not the self-renewal of murine hematopoietic long-term repopulating cells.

The effects of stem cell factor (SCF) have been tested on a murine bone marrow subpopulation (RH123lo, Lin-, Ly6A/E+) that is highly enriched for long-term hematopoietic repopulating cells. SCF maintained cells from this population with long-term repopulating ability for up to 10 days in vitro. However, compared with freshly isolated cells, the level of engraftment in vivo by the cultured cells declined during the in vitro culture period, suggesting that SCF alone was unable to stimulate the self-renewal of long-term repopulating cells. By direct visualization of cultures, only small numbers of cells survived and rarely underwent cell division. However, SCF did directly stimulate proliferation of a population (Rh123med/hi,Lin-,Ly6A/E+) enriched for short-term repopulating cells. These data suggest that stem cell differentiation is associated with the development of mitogenic activity by SCF at least in some progenitor cell populations.

Animals↗
PubMed1994

Induced expression of c-fms in normal hematopoietic cells shows evidence for both conservation and lineage restriction of signal transduction in response to macrophage colony-stimulating factor.

Retrovirus-mediated gene transfer was used to obtain expression of the macrophage colony-stimulating factor (MCSF) receptor, c-fms, on hematopoietic lineages that normally do not express this receptor. Cultures of murine fetal liver cells infected with the c-fms retrovirus developed erythroid colonies in cultures stimulated with M-CSF. However, these colonies were fewer and less hemoglobinized than colonies in parallel cultures stimulated by erythropoietin. Culture of isolated clones demonstrated a direct action of M-CSF on erythroid clones. Culture of c-fms retrovirus-infected adult murine bone marrow cells showed megakaryocyte and novel macrophage-megakaryocyte clones when stimulated by M-CSF. Culture of isolated clones again confirmed a direct action of M-CSF on megakaryocyte clones. In contrast, M-CSF stimulation of c-fms-infected granulocytes and granulocyte progenitor cells did not elicit proliferation, enhanced survival, or functional stimulation of granulocytes. These findings provide evidence for both conservation and lineage restriction of signal transduction in normal hematopoietic cells.

Animals↗
PubMed1994

Design of the abdominal aortic Aneurysm Detection and Management Study. ADAM VA Cooperative Study Group.

PURPOSE: This study describes the design of an ongoing randomized trial intended to determine which of two strategies is superior for managing small abdominal aortic aneurysms (AAA). METHODS: Patients aged 50 to 79 years with AAA 4.0 to 5.4 cm in diameter as determined by computed tomography (CT) who are not at high surgical risk are randomized to either repair of the AAA, called "immediate surgery," or follow-up of the AAA with ultrasonography or CT every 6 months, reserving surgery for those aneurysms that enlarge to 5.5 cm, enlarge rapidly, or become symptomatic, called "selective surgery." RESULTS: The primary outcome measure is all-cause death, and secondary outcome measures are AAA-related death, morbidity, and general health status. The study design calls for 1350 patients to be randomized and monitored for a mean of 5 years. A second objective of the study is to accurately define the prevalence and risk factors for AAA with use of information from the large screening program established to detect AAA for recruitment into the randomized trial. CONCLUSIONS: By the end of 1993, 38,697 patients had been screened with ultrasonography, accounting for about three fourths of new randomizations, and 330 patients had been enrolled (70% of the target rate).

Aged↗
PubMed1994

Frequent immunohistochemical detection of EGF supergene family members in ovarian carcinogenesis.

Primary and metastatic ovarian cystadenocarcinomas, carcinomas of low malignant potential (borderline tumors), benign ovarian cystadenomas, and normal ovaries were compared for immunoperoxidase detection of the ligands epidermal growth factor (EGF), transforming growth factor-alpha (TGF-alpha), amphiregulin (AR), cripto, and the receptors, epidermal growth factor receptor (EGF-R), and c-erbB-2. This matrix analysis of these EGF family members indicated no specific pattern of ligand or receptor expression with a specific ovarian histologic category except in the case of AR and TGF-alpha. AR was detected almost exclusively in borderline tumors, suggesting that these tumors may not arise as a pathological continuum between benign cystadenomas and invasive cystadenocarcinomas. Second, the presence of TGF-alpha immunoreactivity in the absence of coexpression of cripto or EGF appeared to be associated only with adenocarcinomas of high grade and stage.

Amphiregulin↗