Inhibition of the arachidonic acid pathway prevents induction of IL-8 mRNA by phorbol ester and changes the release of IL-8 from HL 60 cells: differential inhibition of induced expression of IL-8, TNF-alpha, IL-1 alpha, and IL-1 beta.

The promyelocytic HL 60 cell line can be used as an in vitro model system to study hematopoetic cell differentiation and inflammatory events. We studied the signal transduction pathway of induced interleukin (IL)-8 expression and compared it with those of tumor necrosis factor alpha (TNF-alpha), IL-1 alpha, and IL-1 beta. The differentiation of HL 60 cells to macrophage-like cells by PMA resulted in a rapid and marked induction of these inflammatory cytokines. The up-regulation occurred in the absence of ongoing protein synthesis, but cycloheximide-sensitive gene products modulated their induction kinetics. Staurosporine, a potent inhibitor of protein kinases, strongly inhibited their gene expression. Phosphorylation may not act directly on latent transcription factors, since bromophenacyl bromide, an inhibitor for the release of arachidonic acid from phorbol-12 myristate 13-acetate (PMA)-stimulated HL 60 cells, markedly depressed the induced mRNAs for IL-8, TNF-alpha, and IL-1 alpha and -beta. Similarly, 5,8,11,14 eicosatetraynoic acid (ETYA), another inhibitor of the arachidonic acid pathway, blocked the induction of transcripts for TNF-alpha, and both IL-1 genes in phorbol ester-stimulated HL 60 cells. In contrast, ETYA increased the induced IL-8 RNA levels and stimulated the release for IL-8. Also, ketoconazole, an inhibitor of 5-lipoxygenase and indomethacin, an inhibitor of cyclooxygenases did not block the induction of IL-8 mRNA. However, the release of IL-8 protein was regulated by indomethacin and ketoconazole. Our results indicate that arachidonic acid metabolites are mediators in the signal transduction pathway of IL-8 expression and that the involved second messengers are different from those which are important for the induction of TNF-alpha and IL-1 beta expression.

The induction kinetics of Il-8 messenger RNA in HL60 cells demonstrate the participation of negative-acting gene(s).

Interleukin-8 (IL-8) mRNA was rapidly, but not permanently, induced at high levels by phorbol-12myristate-13acetate (PMA) in HL60 cells. Ongoing protein synthase does not seem to be required for the initial induction of IL-8 gene expression. However, the rate of transient induction kinetics was modulated by cycloheximide (CHX) indicating that secondary response genes are involved in the regulation of IL-8 RNA levels. Repression of the induced IL8 mRNA by 21 h PMA-treatment was due to reduced transcriptional activity of the gene. In HL60 cells stimulated for 1.5 and 21 h the half-lives of the lL-8 transcripts were markedly increased, suggesting the presence of negatively-acting transcriptional regulator(s).

The differentiation pathway of HL60 cells is a model system for studying the specific regulation of some myeloid genes.

During granulopoiesis, certain myeloid genes encoding products of azurophilic granules are specifically down-regulated. The myeloid specific enzyme myeloperoxidase belongs to this group of genes. It is responsible for the production of hypochlorous acid, a potent microbicidal agent which is involved in host defense. During induced differentiation of promyelocytic leukemic HL60 cells to granulocyte- or monocyte-like cells, myeloperoxidase RNA is depressed. We studied this depression process in more detail by limiting the exposure to the inducer phorbol 12-myristate 13-acetate to 24 h. During this time period, no significant decrease in cell number and cell viability could be observed. Analysis of these in vitro differentiated HL60 cells on the protein and RNA levels showed that they can be used under defined conditions as a cell system to study the specific depression of myeloid genes. Under the described conditions, both the transcriptional rate of the myeloperoxidase gene as well as the stability of its transcript was reduced.

Lyn, a src-like tyrosine-specific protein kinase, is expressed in HL60 cells induced to monocyte-like or granulocyte-like cells.

During the in vitro differentiation of HL60 cells tyrosine-specific kinases are activated. The expression of lyn, a src-related tyrosine kinase, was studied by analysis of the steady-state levels of its transcript during the cell differentiation process induced by retinoic acid, phorbol 12-myristate 13-acetate and 1,25-dihydroxyvitamin D3. In contrast to an earlier report we observe only a small induction of the lyn-RNA levels compared to uninduced control cells. In unstimulated HL60 cells, the level for the lyn-transcript was comparatively high. A second, minor human lyn-transcript with an estimated size of 3.7 kb which has not been previously described, was identified.

Myeloperoxidase is a primary response gene in HL60 cells, directly regulated during hematopoietic differentiation.

The expression of myeloperoxidase was studied in three human myeloid leukemic cell lines. The myeloperoxidase transcript was strongly expressed in promyelocytic HL 60 cells, whereas much lower levels were detected in immature monocytic U 937 cells. Phorbol-12-myristate-13-acetate induction resulted in inhibition of myeloperoxidase expression within 24 hrs. This regulatory event could not be blocked by cycloheximide. Furthermore, cycloheximide did not superinduce myeloperoxidase mRNA levels in KG 1, HL 60 and U 937 cells, arguing against the existence of a negative gene regulator for myeloperoxidase. Therefore, the myeloperoxidase gene can be classified as a primary response gene.

A field portable mass spectrometer for monitoring organic vapors.

A portable mass spectrometer has been designed and built under the sponsorship of the US Army for the purpose of monitoring low concentrations of specified organics in the ambient atmosphere. The goals of the development were discrimination, sensitivity, portability, simplicity of operation, economy and convenience. These objectives were met in a system consisting of a computer operated mass spectrometer with a Llewellyn membrane separator inlet system housed in two 26 x 18 x 9 inch aluminum cases with a total weight less than 150 pounds. This system has shown the capability for field detection of hundreds of specific organic vapors at the parts per billion level in the ambient and workplace environments.

Properties and limitations of hologram recording materials.

The role of the recording material in holographic imaging is reviewed. Following a general classification of holograms into four basic types, the thick and thin amplitude and phase holograms, a discussion is given of the diffraction efficiency and other intrinsic characteristics of each of these types. The effects of zero spread nonlinearities in hologram recording are examined, and the limitations they impose on the usable diffraction efficiency of the various hologram types are discussed. The effect of the recording material modulation transfer function is treated by means of a fictitious mask model which facilitates prediction of the recordable object field and resolution. The noise characteristics of recording media are discussed in terms of their effect on holographic recording. Some examples of noise limitations on holographic information recording by silver halide emulsions are examined, as are the effects of such limitations on the dynamic range of holography. The effects of silver halide grain noise in conventional and holographic recording are compared, showing the conditions under which better performance can be achieved in the latter by a given emulsion. The characteristics of several nonsilver halide hologram recording materials are briefly summarized.