Biomedical subjects

G R Johnson

Publications and source records attributed to G R Johnson.

At least 19 recordsLinked to original sources

PubMed1992

Differential immunohistochemical detection of amphiregulin and cripto in human normal colon and colorectal tumors.

Thirty-six primary human colorectal tumors, 43 noninvolved colon samples that were adjacent to either carcinomas of adenomas, 22 adenomas, and nine normal colon specimens were immunohistochemically examined for the presence and localization of two epidermal growth factor-related peptides, amphiregulin (AR) and cripto. Within the primary tumors, 18 (50%) showed moderate levels of AR expression. Approximately 60% of the tubular and tubulovillous adenomas were positive for AR expression, whereas only 15% of the adjacent, noninvolved colon mucosa expressed AR. A greater proportion of well-differentiated tumors (71%) were positive for AR expression than were poorly differentiated tumors (18%). All of the nine normal colon specimens were positive. Consequently, AR expression appeared to be associated with both normal and malignant epithelial cells that were more differentiated. The distribution of cripto expression was different. Seventy-nine % of the colon tumors expressed cripto with a frequency of expression that was approximately equivalent between well-differentiated and poorly differentiated tumors. Approximately 86% of the tubulovillous adenomas, but only 43% of the tubular adenomas, were positive for cripto expression. In contrast, whereas AR was expressed in normal colon specimens, none of these tissues expressed cripto, and only 12% of the noninvolved normal colon samples adjacent to tumors or adenomas were positive for cripto. Cripto expression therefore appeared related to neoplasia. These data suggest that AR and cripto may be functioning as potential autocrine and/or paracrine growth factors in the colon and that the differential expression of cripto may serve as a potential tumor marker for colonic carcinogenesis.

Adenoma↗

PubMed1992

Rhodamine123 reveals heterogeneity within murine Lin-, Sca-1+ hemopoietic stem cells.

Murine bone marrow Lin-, Ly6A/E+ cells have been fractionated on the basis of rhodamine123 retention into Rh123med/hi and Rh123lo subpopulations. These populations have different responses to hemopoietic growth factors with respect to in vitro colony formation. Cells from either fraction were not stimulated by only granulocyte-colony-stimulating factor (G-CSF), granulocyte/macrophage colony-stimulating factor (GM-CSF), macrophage colony-stimulating factor (M-CSF), interleukins 1 and 6 (IL-1 and -6), or leukemia inhibitory factor (LIF) alone. The Rh123med/hi, but not the Rh123lo fraction, contained cells that could be stimulated by either stem cell factor (SCF) or IL-3 alone. When combinations of growth factors were added, the Rh123med/hi fraction produced more colonies, and responded to a wider range of factor combinations than the Rh123lo population. When tested in vivo, both populations contained no detectable day 8 colony-forming unit-spleen (CFU-S), and similar frequencies of day 13 CFU-S. When transplanted into lethally irradiated recipients (100 cells/recipient), significant numbers of donor cells (67-73%) were found in the peripheral blood of Rh123lo recipients. Both myeloid and lymphoid cells were of donor origin. By comparison, the Rh123med/hi population produced recipients with 1-2% donor cells in peripheral blood, the majority of which were lymphoid.

Animals↗

PubMed1992

Transforming growth factor-alpha acts as an autocrine growth factor in ovarian carcinoma cell lines.

The potential of transforming growth factor-alpha (TGF-alpha) to function as an autocrine growth factor was evaluated in numerous ovarian carcinoma cell lines. All 17 lines which were examined expressed the epidermal growth factor receptor and 16 cell lines, in addition, concomitantly secreted TGF-alpha. Radioimmunoassay of processed serum-free-conditioned medium indicated TGF-alpha concentrations ranging from 16 to 197 pg/ml, or 1.5 to 95 ng/10(8) cells. 125I-TGF-alpha bound to a single class of high-affinity-binding sites on the surface of the cells. The dissociation constant for the 125I-TGF-alpha/epidermal growth factor receptor complex ranged from 0.21 to 5.3 nM with receptor numbers from 3,500 to 96,000/cell, depending upon the cell line. The growth of 8 ovarian cell lines was stimulated in a dose-dependent manner when grown in the presence of exogenous TGF-alpha. Growth in 4 of 5 cell lines capable of serum-free propagation was inhibited from 28 to 56% when cultured in medium containing a TGF-alpha-neutralizing monoclonal antibody. These results support the view that TGF-alpha is an autocrine growth factor for cell lines derived from ovarian cancers of epithelial origin and suggest a potential role for TGF-alpha in the pathogenesis or progression of the disease.

Cell Division↗

PubMed1992

Use of uncalibrated biplanar radiography for the measurement of skeletal coordinates around the shoulder girdle.

Mechanical modelling of the musculoskeletal system is dependent upon information regarding the bony attachments of the relevant muscles; in order to study the biomechanics of the shoulder girdle the authors have identified the muscle attachments in three embalmed cadavers. A simple biplanar radiographic technique was then used to determine the attachment coordinates using frames of reference defined for each bone. This technique, using hand positioning without special fixtures was believed to be sufficiently accurate, bearing in mind the likely degree of biological variation. In order to test this assumption, the accuracy of the technique has been studied by measuring the agreement between the two measurements of the common coordinate in the pairs of radiographs. It was found that for the trunk, the errors in the common coordinate were always less than the natural variation; for the scapula they were of a similar magnitude but, for the humerus, the measurement errors frequently exceeded the variation in the coordinates of muscle attachments. It was concluded that, in general, uncalibrated biplanar radiography was sufficiently accurate for the determination of the spatial coordinates of muscle attachments.

Adult↗

PubMed1992

Autocrine action of amphiregulin in a colon carcinoma cell line and immunocytochemical localization of amphiregulin in human colon.

Amphiregulin (AR) is a newly discovered glycosylated, 84-amino acid residue polypeptide growth regulator which has sequence homology to the EGF family of proteins. To obtain immunological reagents to study the biological role of AR, two synthetic peptides containing sequences corresponding to distinct regions of AR were used to generate polyclonal antibodies in rabbits. One preparation of antipeptide antibodies directed against residues 26-44 of AR (AR-Ab2) was most effective in the detection of native AR, whereas another preparation of antibodies against residues 8-26 (AR-Ab1) was found to be most efficacious in the detection of AR in formalin-fixed and paraffin-embedded tissues. The growth of a colon carcinoma cell line, Geo, which proliferates autonomously under serum-free conditions, was stimulated by the exogenous addition of AR or EGF. Half-maximal stimulation of this growth was observed at 40 and 200 pM of EGF and AR, respectively. A mAb to the extracellular domain of the EGF receptor blocked the stimulation of cell proliferation induced by the exogenous addition of AR, suggesting that this stimulation was mediated via the EGF receptor. Geo cells were found to constitutively express significant levels of the AR mRNA transcript as determined by analysis of the polymerase chain reaction-amplified cDNA product and AR protein was detected immunocytochemically using the AR-Ab1 antibodies in these cells. AR was immunoprecipitated specifically using the AR-Ab2 antibodies from the conditioned medium of Geo cells, which had been metabolically labeled with [35S]cysteine. The secreted AR migrated as a broad band (18.5-22.5 kD) with a median molecular weight of approximately 20.7 kD in SDS-PAGE. Immunospecific removal of AR from serum-free medium conditioned by the Geo cells and readdition of the AR-depleted medium to Geo cells resulted in an approximately 40% inhibition of cell growth relative to controls. Furthermore, the growth of the Geo cells was also inhibited by approximately 50% by the addition of the anti-EGF receptor mAb alone. These results indicate that AR and the EGF receptor are involved in the autocrine growth of these cells and suggests that AR may act through the EGF receptor via an extracellular autocrine loop. To study the expression of AR in human colon in vivo, AR was localized immunocytochemically in formalin-fixed, paraffin-embedded sections from normal and malignant human colon using the AR-Ab1 antibodies.(ABSTRACT TRUNCATED AT 400 WORDS)

Amphiregulin↗

PubMed1992

Tissue expansion as an alternative to skin grafting for closure of skin deficits.

Recent advances in the technical aspects of tissue expanders for closure of skin deficits have led to an overall reduction in the high rate of complications typically associated with their usage. The authors provide a review of the recent refinements in the technique of the tissue expansion process. A discussion of current and future applications using this technique in the treatment of various lower extremity pathologies, including clubfoot, is presented. A case report on the use of a subcutaneous tissue expander for the treatment of a failed skin graft of the medial ankle is included.

Adult↗

PubMed1992

Long-term hemopoietic repopulation by Thy-1lo, Lin-, Ly6A/E+ cells.

Murine bone marrow Thy-1lo, Lin-, Ly6A/E+ cells have been isolated by fluorescence-activated cell sorting (FACS). This population contained up to 27% in vitro colony-forming cells (CFC) when stimulated by interleukin 3 (IL-3) and 5% day-12 spleen colony-forming units (CFU-S) (uncorrected for seeding efficiency). As few as 100 cells were able to reconstitute the myeloid and lymphoid compartments of lethally irradiated recipients for periods of up to 72 weeks. Over the range of 30-560 transplanted cells, three patterns of engraftment were observed; the proportion of donor cells increased gradually to values of > 90%, gradually declined, or remained static over the 72-week observation period. When the donor cell percentage in the peripheral blood exceeded 60%, both myeloid (neutrophils and monocytes) and lymphoid (T- and B-lymphocytes) cells were of donor origin. Primary animals containing > 60% donor cells in their peripheral blood were able to engraft secondary recipients with donor cells of both myeloid and lymphoid lineages. Primary animals with < 20% donor cells in their peripheral blood contained bone marrow cells that were only able to produce lymphoid cells in the peripheral blood of secondary recipients. With > 90% of animals, including both primary and secondary recipients, there were no large fluctuations in the proportion of donor myeloid or lymphoid cells in the peripheral blood. Where changes occurred, with three exceptions, these were gradual. These data suggest that the Thy-1lo, Lin-, Ly6A/E+ population is heterogeneous and contains in vitro CFC, day-12 CFU-S, and long-term repopulating cells for both the myeloid and lymphoid lineages and cells capable of long-term repopulation of only the lymphoid lineage.

Animals↗

PubMed1992

Fatal polycythemia induced in mice by dysregulated erythropoietin production by hematopoietic cells.

C57BL/6J murine bone marrow cells, infected with a retroviral vector (MP Zen) carrying a monkey erythropoietin cDNA, were transplanted into lethally irradiated syngeneic recipients to study the effect of erythropoietin production by hemopoietic cells. High levels of erythropoietin were recorded in the plasma (median value: 1.2 u/ml) and in media conditioned by peritoneal, spleen, and bone marrow cells from recipient mice. In transplanted mice, the hematocrit was elevated (90 +/- 5%) and the mice died at a mean of 71 days after transplantation. In the blood, platelet counts were usually low and nucleated blood cells slightly elevated. Spleen weight increased 5-fold and bone marrow cellularity decreased slightly. There was a 9.9-fold increase in erythroblast numbers, a 2-fold reduction of lymphocytes, and no variation of the myeloid cells when the total cellularity of bone marrow, spleen, peripheral blood, and peritoneal cells were considered. Calculation of the total numbers of progenitor cells in these organs revealed a 18-fold increase in erythroid colony-forming units (CFU-E) but no significant variation of the erythroid burst-forming units (BFU-E), and myeloid progenitor cell numbers. A variable proportion of CFU-E, (12% or 24% in bone marrow or spleen, respectively) was able to proliferate in unstimulated cultures. Erythropoietic amplification occurred in the spleen and there was a redistribution of the BFU-E and myeloid cells from the bone marrow to the spleen. No significant extramedullary erythropoiesis was seen. This study emphasizes the erythroid specificity of erythropoietin and shows that elevated dysregulated erythropoietin production by hemopoietic cells leads to a fatal polycythemia without erythroid neoplastic transformation.

Animals↗

PubMed1991

Developmental potential of the earliest precursor cells from the adult mouse thymus.

A new, numerically minute population of cells representing the earliest T precursor cells in the adult mouse thymus has recently been isolated. This population has been shown to be similar to bone marrow hemopoietic stem cells in surface antigenic phenotype and to express moderate levels of CD4. We now show, by fluorescence-activated cell sorting and intrathymic transfer to irradiated mice, that this apparently homogeneous population differs from multipotent stem cells in expressing the surface stem cell antigen 2 (Sca-2), that it differs from most early B lineage cells in lacking B220 and class II major histocompatibility complex expression, and that it binds rhodamine 123 like an activated rather than a quiescent cell. Irradiated recipient mice differing at the Ly 5 locus were used to compare the developmental potential of these early intrathymic precursors with bone marrow stem cells. Only T lineage product cells were detected when the intrathymic precursor population was transferred back into an irradiated thymus. However, when the intrathymic precursor population was transferred intravenously, it displayed the capacity to develop into both B and T lymphoid cells in recipient bone marrow, spleen, and lymph nodes, but no donor-derived myeloid cells were detected. The absence of myeloid and erythroid precursor activity was confirmed by showing that the intrathymic precursor population was unable to develop into myeloid or erythroid spleen colonies on intravenous transfer or to form colonies in an agar culture. These findings indicate that this earliest intrathymic precursor population has become restricted (or strongly biased) to lymphoid lineage development, but not exclusively to T lymphocytes.

Agar↗

PubMed1991

Response to and expression of amphiregulin by ovarian carcinoma and normal ovarian surface epithelial cells: nuclear localization of endogenous amphiregulin.

Amphiregulin (AR) is a polypeptide growth regulator which has sequence homology to the epidermal growth factor-related family of ligands and contains putative nuclear targeting sequences. Human ovarian carcinoma cell lines and their normal counterparts, ovarian surface epithelial cells (OSEs), were assessed for their ability to respond to and express AR. Addition of exogenous AR (8-200 pM) inhibited the growth of 2 of 3 OSE specimens and 3 of the 6 carcinoma cell lines indicating that AR has the potential to inhibit the growth of normal cells, in addition to carcinoma cells. In contrast, concentrations of AR ranging from 1-5 nM stimulated the growth of all 3 of the OSEs and 4 of the 6 carcinoma cell lines. Immunocytochemical staining of the cells using antipeptide antibodies directed against residues 8-26 of AR indicated that all cells expressed AR and that the staining was localized to the nucleus. The nuclear staining of AR was concentrated in the nucleolus of the carcinoma cells, whereas the staining was diffuse in the nucleus of the OSEs. These results suggest that AR may play a growth regulatory role in the nucleus of cells and this role may be different in normal and malignant epithelial cells.

Amphiregulin↗

PubMed1991

Ranges of movement at the shoulder complex using an electromagnetic movement sensor.

There are few published data on the ranges of available motion at the shoulder and no clinical technique for the assessment of three dimensional movement. An electromagnetic movement sensor (Isotrak), which makes three dimensional measurements of shoulder movement and measures the ranges of flexion, extension, abduction in the neutral position, abduction preceded by external rotation, and internal rotation achieved by placing the hand as far up the back as possible, was used to study 16 normal subjects. Intraobserver repeatability was good, and a study of five subjects by two observers showed no significant difference between single measurements made by each. The measured ranges were in broad agreement with published data, though they tended to be smaller. This was attributed to either the bulk of the mounting for the movement sensor or to the immobilisation of the elbow and hence the biceps muscle. Women had greater ranges, except for externally rotated abduction.

Analysis of Variance↗

PubMed1991

Effects on spleen colony-forming unit self-renewal after retroviral-mediated gene transfer of multi-colony-stimulating factor, granulocyte-macrophage colony-stimulating factor, or granulocyte colony-stimulating factor.

C57BL/6 mice were established to constitutively produce multi-colony-stimulating factor (multi-CSF), granulocyte-macrophage colony-stimulating factor, or granulocyte colony-stimulating factor by transplantation with post-5-fluorouracil-treated syngeneic marrow cells infected with retroviral vectors bearing the corresponding growth factor complementary DNAs. At 2-4 weeks after transplantation, these mice had a 2-7-fold increase in spleen colony-forming unit (CFU-S) numbers when compared to control animals transplanted with MPZipNeo-infected marrow cells. Most of the CFU-S in the former animals were located in the spleen, whereas those of control mice were found in the marrow. The increase in total CFU-S content in recipients of CSF-infected mice was not due to an increase in self-renewal ability but rather to the recruitment of more primitive cells, as there were no differences in CFU-S content of spleen colonies generated from marrow cells of either group. Neither were there any differences in the cellular composition of these spleen colonies or in the size or distribution of cell types in secondary spleen colonies generated from these spleen colonies, suggesting the inability of any of the three CSFs to alter the differentiation pattern of CFU-S.

Animals↗

PubMed1991

Comparison of two instructional methods on the levels of physiologic and psychologic stress as measured by blood cortisol and the State-Trait Anxiety Inventory.

This study was designed to determine whether a systematic or nonsystematic instructional strategy affected the levels of physiologic and psychologic stress as measured by blood cortisol levels and the State-Trait Anxiety Inventory (STAI) in students in the postsecondary educational setting. A convenience sample consisting of 43 subjects was randomly assigned to either a systematic or nonsystematic teaching group. The blood cortisol and STAI were measured 1 and 2 weeks before the treatment and following the treatment on the day of the study. Results of the study demonstrated that there were differential posttreatment increases in the amount of physiologic stress, as measured by blood cortisol levels produced by either instructional method. However, between the control measurement 1 week before the treatment and the posttreatment measurement, there were no effects observed for the psychologic STAI measures for either group. Accounting for the circadian rhythm effect of cortisol, there was a significant "buffering effect" in stress experienced by the subjects in the systematic teaching group. More specifically, the nonsystematic teaching group experienced a 55.42% rate increase in cortisol compared to a 10.74% rate increase for the systematic teaching group which was statistically significant. The systematic teaching method may be more effective in preventing physiologic stress in the educational setting and possibly in the clinical practice of anesthesia nursing. Additionally, the results suggested that the STAI may be inappropriate when used as an index of stress in certain educational settings.

Circadian Rhythm↗

PubMed1990

Rapid receptor-mediated catabolism of 125I-atrial natriuretic factor by vascular endothelial cells.

The binding, internalization and degradation of 200 pM monoiodinated human atrial natriuretic factor-(99-126) (125I-hANF) by cultured bovine aortic endothelial cells (BAECs) were studied at 37 degrees C. 125I-hANF was rapidly cleared from the extracellular medium (t1/2 approximately 10 min), whereas preincubation of the cells in the presence of 20 mM-NH4Cl or 0.2 mM-chloroquine resulted in a significant inhibition of this process. The BAECs rapidly produce three major degradation products of 125I-hANF, namely [125I]iodotyrosine 126 (125I-Y), Arg125-[125I]iodotyrosine126 (125I-RY) and Phe124-Arg125-[125I]iodotyrosine126(125I-FRY), which were detected in the extracellular medium. NH4Cl and chloroquine acted to inhibit the generation of 125I-Y and 125I-RY, but not that of 125I-FRY. Furthermore, excess unlabelled hANF (300 nM) completely blocked the rapid production of 125I-Y and 125I-RY in the first 5 min, but only partially (49%) inhibited the generation of 125I-FRY. Thus, in contrast with our previous findings with cultured smooth-muscle cells [Johnson, Arik & Foster (1989) J. Biol. Chem. 264, 11637-11642], BAECs bind, internalize and rapidly degrade 125I-hANF, resulting in the release of 125I-Y and 125I-RY into the extracellular medium. Similarly to smooth-muscle cells, the BAECs generate 125I-FRY from 125I-hANF via an extracellular proteolytic event. The rapidity of the receptor-mediated process and its sensitivity to NH4Cl and chloroquine suggest that the 125I-hANF is proteolytically processed in the endosomes of BAECs and that its receptors cycle between the cell surface and intracellular stores.

Ammonium Chloride↗

PubMed1990

The complete sequence of the acidic subunit from Mojave toxin determined by Edman degradation and mass spectrometry.

Mojave toxin, a heterodimeric, neurotoxic phospholipase complex from Crotalus scutulatus scutulatus, is one of a group of closely related rattlesnake toxins for which much structural information is still lacking. The complete amino-acid sequence of the acidic subunit from Mojave toxin was determined. The three individual peptide chains, derived from the acidic subunit by reductive alkylation, were separated by high-performance liquid chromatography. Fragmentations of the A and B chains were done using specific proteinases and the resulting peptide mixtures were fractionated by reverse-phase high-performance liquid chromatography. Sequence analyses on the intact chains and the fragments from digests were done by automated Edman degradation, carboxypeptidase Y degradation and triple-quadrupole and tandem-quadrupole Fourier-transform mass spectrometry. The sequence for each acidic subunit chain is very similar to the corresponding chain from the related neurotoxin complex, crotoxin, and overall the sequence is similar to the sequences of group I and II phospholipases A2. The N-terminus of the B chain is blocked by pyroglutamic acid. The existence of two distinct and closely related C chains was established. It is unlikely that the small sequence difference can account for the isoforms that are present in purified Mojave toxin and in unfractionated venom.

Amino Acid Sequence↗

PubMed1990

Partial characterization of a metalloendopeptidase activity produced by cultured endothelial cells that removes the COOH-terminal tripeptide from 125I-atrial natriuretic factor.

The presence of the COOH-terminal region of human atrial natriuretic factor-(99-126) (hANF) is necessary for the full expression of its biological activity. Here, we report on the partial characterization of a proteolytic activity in the conditioned medium from cultured bovine aortic endothelial cells that cleaves the Ser123-Phe124 bond of 125I-hANF generating the COOH-terminal tripeptide. The concentrated conditioned medium was fractionated by gel filtration high performance liquid chromatography and fractions were assayed for the ability to generate the COOH-terminal tripeptide from 125I-hANF. This analysis indicated that the protein responsible for this activity had an approximate molecular weight of 200,000 daltons. Of 16 protease inhibitors tested, only 1,10 phenanthroline, EDTA, EGTA and N-ethylmaleimide significantly inhibited the endopeptidase activity. Thus, we conclude that cultured bovine aortic endothelial cells produce a potentially novel phosphoramidon-insensitive metalloendopeptidase that removes the COOH-terminal tripeptide from 125I-hANF.

Animals↗

PubMed1990

Proliferative properties of unfractionated, purified, and single cell human progenitor populations stimulated by recombinant human interleukin-3.

In this report, the biological properties of human recombinant interleukin-3 (rhIL-3) were studied. We investigated the range of unfractionated, purified and single cell human progenitors responsive to IL-3; compared the colony types observed with those obtained in the presence of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) or granulocyte-CSF (G-CSF). The results show that IL-3 directly stimulates the formation of colonies derived from eosinophil and, to a lesser degree, granulocyte and macrophage progenitors. In combination with erythropoietin, it supports the development of erythroid and mixed-erythroid colonies. Furthermore, the data show that IL-3 is a more potent stimulus for both erythroid and eosinophil progenitors than GM-CSF. Interleukin-3 stimulates the formation of both compact and dispersed colonies derived from eosinophil progenitors, whereas GM-CSF stimulates the formation of only the compact type. We conclude that some of the proliferative effects of IL-3 observed on unfractionated and semipurified bone marrow populations are indirect and most likely involve accessory cell interactions.

Bone Marrow Cells↗

PubMed1990

Relationship between domoic acid levels in the blue mussel (Mytilus edulis) and toxicity in mice.

Monitoring of eastern blue mussels (Mytilus edulis), contaminated with domoic acid, involved mouse bioassays and quantitative analysis using HPLC. Mice undergo a typical scratching syndrome at sublethal as well as lethal doses of domoic acid. The onset of scratching behaviour and time of death in mice were inversely related to the dosage of domoic acid. An LD50 (i.p.) of 3.6 mg domoic acid/kg mouse was calculated. Toxic mussels held in tanks and flushed with uncontaminated sea water showed a decline in domoic acid concentration in mussel tissue with time. In addition, domoic acid concentrations in mussels from two infected rivers declined to negligible levels in 40-50 days under normal environmental conditions. The bulk of domoic acid and toxicity was located in the hepatopancreas which also contained large amounts of chlorophyll-A, an algae biomass indicator, relative to control mussels. These results support the conclusion that domoic acid was the primary causative factor in the shellfish poisonings from Prince Edward Island mussels in late 1987.

Animals↗