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J M Lord

Publications and source records attributed to J M Lord.

At least 145 records · Page 8Linked to original sources
PubMed1989

Evidence that precursor cells of monocytes and B-lymphocytes are closely related.

Various data in the literature suggest that progenitor cells of monocytes and B cells are closely related. In this study we have investigated this notion by using two-dimensional gel electrophoresis patterns of total cellular phosphoproteins to assess the similarity and thus the close relationships between erythroid, granulocyte, monocyte, and B-lymphocyte cell lines that typify immature cells of these lineages. In previous studies, six proteins had been identified whose constitutive phosphorylation correlated with the capacity of HL60 variant cell lines to differentiate towards either neutrophils (four proteins) or monocytes (two proteins). The presence or absence of five of these phosphoproteins in autoradiographs obtained for the pre-B-cell lines Nalm6 and SMSB showed that the pre-B-cell lines most closely resembled lines able to differentiate towards monocytes (HL6015-12, HL60M2, U937, and ML-1) as opposed to lines restricted to neutrophil (HL60Ast3) or erythroid differentiation (K562). The K562 phosphoprotein pattern resembled that obtained for HL60Ast3. Progressive changes in the constitutive phosphorylation of the five proteins were observed suggesting that cells that have acquired the potential for either neutrophil, monocyte, or B-cell differentiation have initially diversified in a linear progressive manner. This observation supports a model for hemopoiesis that suggests that, during progenitor cell development, differentiation potentials are expressed individually in the above order. Two additional phosphoprotein spots were found to be restricted to the pre-B-cell lines. These phosphoproteins, together with those that change their intensity in autoradiographs of erythroid, myeloid, and B-cell lines, suggest that protein phosphorylation plays an important role during cell diversification.

B-Lymphocytes↗
PubMed1988

A rapid procedure for isolating hemopoietic cell nuclei.

A new method for isolating cell nuclei is described which involves freezing and thawing cells in 2% Tween 40, then gentle homogenization to release nuclei, followed by immediate microcentrifugation through 50% sucrose. Purified nuclei were obtained in 3 min and yields of 78-95% were obtained from a variety of human hemopoietic cells. Electron microscope analysis of nuclei obtained from HL60 cells showed that 89% of the nuclei were intact and have an appropriate morphology. A low level of contamination with other organelles was revealed by electron microscopy and by using specific assays for plasma membrane, mitochondria, lysosomes, Golgi membrane, and endoplasmic reticulum (0.5-5.5%). The value of the technique is that nuclear proteins and small metabolites which might be lost by rapid leakage from isolated nuclei and the possibility of biochemical modification of cellular constituents are minimized by using a rapid isolation procedure.

Cell Fractionation↗
PubMed1988

The expression of functional ricin B-chain in Saccharomyces cerevisiae.

Yeast cells transformed with plasmids containing ricin B-chain coding sequences expressed this heterologous protein. When ricin B-chain was expressed in a form which resulted in its deposition in the yeast cytosol it formed insoluble aggregates which were devoid of galactose-binding activity. In contrast, when DNA fusions were constructed, in which the B-chain coding sequence was preceded by either the preproalpha-factor leader sequence or the native preproricin signal sequence, the recombinant B-chain products were soluble and biologically active. Both the homologous yeast signal peptide and the heterologous plant signal peptide directed the expressed product into the lumen of the yeast endoplasmic reticulum. As a result, the recombinant B-chain products were processed at the N-terminus, glycosylated and folded into an active conformation, presumably stabilized by correct intrachain disulphide bond formation.

Base Sequence↗
PubMed1988

Near neighbour analysis of variant cell lines derived from the promyeloid cell line HL60.

The human promyeloid cell line H60 can be induced to differentiate towards either neutrophils or monocytes. Variant cell lines, derived from HL60, which show reduced capacities for neutrophil and monocyte differentiation can be arranged in a developmental sequence which suggests that the potentials for neutrophil and monocyte differentiation are expressed sequentially by HL60 cells in this order. Analysis of the patterns of total cellular phosphoproteins within HL60 and 5 variant cell lines, by two-dimensional gel electrophoresis, has identified 6 distinct phosphoproteins which show progressive differences in the intensity of spots between the variant lines. The changes in these phosphoproteins relate to the position of the lines within the proposed development sequence. Similarly, lines placed close together in the sequence are more similar, as regards phosphoprotein profiles, than lines placed far apart. These studies provide direct evidence in favour of the hypothesis that the potentials for neutrophil and monocyte differentiation are expressed sequentially during myelopoiesis. Furthermore, two phosphoprotein spots were found to be restricted to lines able to differentiate towards monocytes. These proteins may play important roles during commitment to monocyte differentiation.

Bone Marrow Cells↗
PubMed1988

The development of cell lineages: a sequential model.

The concept of cell lineage and the empirical characterization of specific lineages provide valuable insight into the problems of developmental biology. Of central interest is the decision-making process that results in the diversification of cell lines. Studies of the haemopoietic system, in which stem cells can be committed to one of at least six pathways of differentiation, have suggested that the restriction of differentiation potentials is a progressive and stochastic process. We have recently proposed an alternative model which hypothesizes that lineage potentials during haemopoiesis are expressed individually and in a predetermined sequence as progenitor cells mature. The model first arises from experimental studies which show that both normal myeloid progenitor cells and a human promyeloid cell line, which are able to differentiate towards either neutrophils or monocytes, express these potentials sequentially in culture. The close linear relationship between other haemopoietic progenitor cells is inferred from collective data from studies of bipotent progenitor cells and of haemopoietic proliferative disorders. If the development of haemopoietic cell lineages shows a tendency to follow a particular program, such a mechanism is likely to operate throughout development. In this paper we consider the evidence in favour of programmed events within progenitor cells implementing diversification, and the implications of predetermined and restricted pathways of embryonic development.

Animals↗
PubMed1988

Characterization of a novel nuclear envelope protein restricted to certain cell types.

The monoclonal antibody AGF2.3 identifies a nuclear envelope protein that is restricted to certain cell types. In particular, this antigen shows a reduced level of expression during haemopoietic cell maturation. In this study, we have examined the relationship of this protein to known nuclear envelope proteins that have a similar molecular mass. Antigen extraction and immunoelectron microscope studies revealed that the AGF2.3 protein is an integral membrane protein present at both the inner and outer aspects of the nuclear envelope. The protein is not associated with nuclear pores and therefore is distinct from pore complex proteins. The AGF2.3 protein does not have ATPase activity. Therefore, this protein is also distinct from a myosin heavy chain-like ATPase that is associated with the nuclear envelope. The AGF2.3 antibody identifies a novel nuclear envelope protein. Further studies of the biochemical nature of the AGF2.3 protein should provide insight into novel cellular processes at the nuclear envelope relating to the lineage or maturation status of cells.

Antibodies, Monoclonal↗
PubMed1988

Changes in insulin receptor expression in HL60 cells induced to differentiate towards neutrophils or monocytes.

HL60 is a human promyeloid cell line capable of differentiating towards monocytes or granulocytes when treated with appropriate agents. Changes in insulin receptor number, affinity and mRNA levels were observed when HL60 cells were induced to differentiate with 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or dimethylsulphoxide (DMSO). Total and high-affinity insulin receptor numbers decreased following treatment of HL60 cells with DMSO, whereas total insulin receptor number increased and high-affinity receptor number decreased in cells treated with TPA. Three distinct insulin receptor mRNA species of 9.1, 6.3 and 2.8 kb were identified in HL60 cells. The larger 9.1 and 6.3 kb species were increased in both TPA- and DMSO-treated HL60 cells, and the 2.8 kb mRNA was reduced in differentiated cells. Thus HL60 cells differentiated towards monocytes or granulocytes showed similar changes in the levels of individual insulin receptor mRNAs, but displayed contrasting alterations in low-affinity insulin binding. Three HL60 variant lines, which have different capacities to respond to inducers of monocyte and neutrophil differentiation, showed similar levels of total insulin receptors, but differed in their expression of high-affinity receptors. The data provide evidence for the existence of two distinct insulin receptors.

Blotting, Northern↗
PubMed1988

Changes in phosphoproteins during commitment of HL60 cells to monocyte differentiation: evidence for multiple protein kinase involvement.

The human promyeloid cell line HL60 differentiates toward monocytes when treated with TPA. We have analyzed, by two-dimensional gel electrophoresis, the phosphoprotein patterns within HL60 cells, labeled to equilibrium with [32P]orthophosphate when cells were treated with suboptimal (1 nM), optimal (5 and 10 nM), and supraoptimal (40 and 100 nM) concentrations of 12-O-tetradecanylphorbol-13-acetate (TPA) as regards the induction of differentiation. No change was detected in the phosphoprotein pattern at 1 nM TPA, whereas four phosphoproteins showed increased levels of phosphorylation at 5 and 10 nM TPA. When cells were treated with 40 and 100 nM TPA, in total eight and ten proteins, respectively, were phosphorylated, including the above four proteins. Two proteins were dephosphorylated when cells were treated with 40 and 100 nM TPA. A 15-kd protein, phosphorylated when HL60 cells were treated with 5 nM TPA, was observed as an intense spot in autoradiographs of total cellular phosphoproteins of two variant HL60 cell lines that are unable to differentiate toward monocytes and prior to treatment with TPA. In the case of three variant cell lines, which like HL60 differentiate toward monocytes, the phosphoprotein spot was almost absent. Thus, paradoxically, the 15-kd phosphoprotein is affected by TPA although its constitutive level of expression or increased phosphorylation state is inversely related to the potential for monocyte differentiation. This observation, together with the TPA dose-response effects on protein phosphorylation, is discussed in relation to multiple protein kinase involvement.

Cell Differentiation↗
PubMed1987

Expression of ricin A chain in Escherichia coli.

DNA encoding ricin A chain was derived from preproricin cDNA and ligated into the expression vector pDS5/3. Transcription is controlled from the coliphage promoter PN25 fused with the lac operator of E.coli. When induced, E.coli 71.18 cells transformed with the recombinant plasmid express ricin A chain which is soluble and has full biological activity.

DNA↗
PubMed1987

The use of cytotoxic plant lectins in cancer therapy.

As part of their defense mechanism against herbivores or phytophagous insects, many plant tissues contain lectins. Some of these lectins are potent toxins which kill animal cells by arresting protein synthesis. An attractive strategy for developing specifically cytotoxic chemotherapeutic agents is to link cell type-specific monoclonal antibodies to potent toxins. The plant protein ricin has emerged as the toxin of choice for such constructs.

Journal Article↗
PubMed1987

Brain stem evoked response in the assessment of learning disabilities.

Evidence has accumulated rapidly to support the hypothesis that neurological anomalies underlie specific forms of learning disability (LD). This article reviews recent studies directed at the electrophysiological assessment of LD children, with emphasis on the brain stem evoked response(BSER) methodology. Child studies appear to indicate that there are significant differences in BSERs between LD and age-matched normal controls. The methodological and statistical differences among these studies, which have clouded the findings and made replication difficult, are reviewed. Improvements in sampling and empirical procedures are suggested, including the use of discriminant analysis to explore BSER asymmetries in LD children. Future inquiry into the measurement and analysis of BSER activity in LD populations is warranted on the basis of these preliminary findings. New information may bring professionals closer to an understanding of the neurological causes of specific subtypes of LD and permit earlier identification of these "at risk" populations.

Adolescent↗
PubMed1987

Plant toxins.

Many plants produce toxic proteins capable of inactivating eukaryotic ribosomes and thereby arresting protein synthesis. In certain plants the gene for a ribosome-inactivating protein has fused with a gene encoding an independent sugar-binding polypeptide to generate heterodimeric proteins which are among the most potent cytotoxins known.

Cytotoxins↗