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Biomedical subjects

M Herzberg

Publications and source records attributed to M Herzberg.

At least 19 recordsLinked to original sources
PubMed1998

Platelet-derived interleukin-1 induces cytokine production, but not proliferation of human vascular smooth muscle cells.

During vascular injury, such as observed in atherosclerosis, restenosis, vasculitides, transplantation, or sepsis, vascular smooth muscle cells (SMC) can be exposed to platelets or platelet products. Under these conditions proliferation or cytokine production of SMC stimulated by platelets or platelet products may contribute to regulation of vascular pathogenesis. Thus, we investigated interleukin-6 (IL-6) and IL-8 production as well as proliferation of SMC in response to platelets or platelet lysates. Platelets not already preactivated by thrombin induced IL-6 (10- to 50-fold) or IL-8 production of unstimulated SMC in a cell number dependent fashion. Preactivation of platelets with thrombin potently increased the platelet-mediated IL-6 (50- to 1,000-fold) and IL-8 production of SMC. Hirudin specifically inhibited the activation of platelets with thrombin. Isolated platelets cultured in the absence of SMC did not contain detectable IL-6 or IL-8. Prestimulation (4 hours) of SMC with pathophysiologically relevant substances (lipopolysaccharide [LPS], tumor necrosis factor-alpha [TNF-alpha], or IL-1alpha) further increased the platelet-induced cytokine production. The platelet-derived SMC stimulatory activity was IL-1, since IL-1 receptor antagonist (IL-1-Ra) inhibited the platelet-induced cytokine production of SMC. Anti-platelet-derived growth factor (PDGF)-antibody did not further reduce this activity. Thrombin itself stimulated expression of IL-6 and IL-8 to some degree and induced IL-6 production of SMC synergistically with IL-1. Platelets also induced proliferation of SMC, however, anti-PDGF antibodies, rather than IL-1-Ra blocked this response. These data show that platelet-derived IL-1 stimulates cytokine production of vascular smooth muscle cells, indicating that platelet-derived IL-1 may contribute to regulation of local pathogenesis in the vessel wall by activation of the cytokine regulatory network.

Antibodies, Monoclonal↗
PubMed1997

Human vascular smooth muscle cells express interleukin-1beta-converting enzyme (ICE), but inhibit processing of the interleukin-1beta precursor by ICE.

Local immunoregulatory processes during normal vascular biology or pathogenesis are mediated in part by the production of and response to cytokines by vessel wall cells. Among these cytokines interleukin (IL)-1 is considered to be of major importance. Although vascular smooth muscle (SMC) and endothelial cells (EC) expressed both IL-1alpha and IL-1beta as cell-associated, 33-kilodalton (kD) precursors, SMC neither contained detectable mature IL-1beta, nor processed recombinant IL-1beta precursor into its mature 17-kD form. Thus, we investigated the expression and function of IL-1beta-converting enzyme (ICE) in vascular cells. We demonstrate in processing experiments with recombinant IL-1 precursor molecules that EC processed IL-1beta, in contrast to SMC. Despite the failure of SMC to process IL-1beta, these cells expressed ICE mRNA, immunoreactive ICE protein, and the expected IL-1beta nucleotide sequence. The lack of processing was explained by our finding that extracts of SMC specifically and concentration dependently blocked processing of IL-1beta precursor by recombinant or native ICE. The initial biochemical characterization of the inhibitory activity showed that it is heat-labile, has a molecular size of 50-100 kD, and is associated to the cell membrane compartment. Inhibition of processing, i.e., activation of IL-1beta precursor by SMC may constitute a novel regulatory mechanism during normal vascular biology or pathogenesis of vascular diseases.

Caspase 1↗
PubMed1997

Characterization of prosomes in human lymphocyte subpopulations and their presence as surface antigens.

Prosomes, also called "multicatalytic proteinase" (MCP) or "proteasomes," are a new type of ubiquitous RNP particle present in some archeobacteria and in all eukaryotic cells tested from yeast to human. They were discovered as subcomplexes of untranslated messenger-ribonucleoproteins (mRNP) and later found to have a MCP activity putatively involved in antigen processing. Being composed of variable sets of characteristic proteins and associating small RNAs (pRNA), families of individual "mosaic" prosome particles seem to characterize the differentiation type and physiological state of individual cells and tissues. Here, prosomes from human lymphocytes, isolated and characterized biochemically and by Western blot analysis, were found to differ in their subunit composition compared to other human prosomes. Surprisingly, prosomal antigens were discovered at the outer surface of blood cells monitored by flow cytometry with monoclonal antibodies to individual prosomal proteins. It was observed that human T and B lymphocytes have variable and characteristic prosomal antigens at their surface according to their CD classification. Interestingly, the lymphocyte subpopulations most strongly labeled by the anti-p25K and anti-p27K mAbs were the NK and B cells.

Adult↗
PubMed1996

The effect of estrogen replacement therapy on zinc in serum and urine.

OBJECTIVE: To ascertain the influence of estrogen replacement therapy (ERT) on blood and urinary zinc in postmenopausal women. METHODS: Thirty-seven postmenopausal women aged 53.2 +/- 3.7 years were examined. All were treated with conjugated estrogens 0.625 mg and medroxyprogesterone acetate 5 mg. Zinc, magnesium, calcium, phosphate, and alkaline phosphatase levels in blood were measured before and after 6 and 12 months of treatment. Urinary excretion of zinc, magnesium, calcium, phosphate, and hydroxyproline were evaluated before and after 3, 6, and 12 months of therapy. Bone mineral density was examined before treatment and after 1.7 +/- 0.3 years of ERT. Subjects were classified by 1) initial bone mineral density values (osteoporotics less than 0.850 g/cm2) and 2) zinc excretion as elevated (greater than 600 micrograms/g creatinine). RESULTS: At baseline, the values of most markers of bone turnover were higher in the osteoporotic women (Hotelling test, P = .06). After 1 year of treatment, a higher decrease of most indices was observed in the osteoporotic patients, and no statistical difference was found between the osteoporotic and the normal groups (Hotelling test, P = .31). A consistent negative association was observed between changes in bone mineral density and urinary zinc excretion in the osteoporosis group. Estrogen replacement therapy reduced excretion of zinc, magnesium, and hydroxyproline in the elevated zinc excretion group. Zinc excretion decreased 35% after 3 months and 26% after 1 year of treatment. The serum tests, with the exception of alkaline phosphatase, showed only negligible changes during ERT. CONCLUSION: A significant decrease in zinc excretion was observed after 3 months of ERT. This change was more pronounced in women with osteoporosis and elevated zinc excretion. Because zinc excretion is almost uninfluenced by variation in diet, it may be used as an additional marker of changes in bone metabolism.

Bone Density↗
PubMed1993

A novel rapid hybridization technique: paper chromatography hybridization assay (PACHA).

A new DNA hybridization technique, based on chromatographic migration of DNA on a nitrocellulose strip passing through an immobilized probe area, is described. The new paper chromatography hybridization assay (PACHA) is faster and simpler to use than the conventional dot hybridization assay. In this assay, an aliquot of biotinylated, PCR-amplified target DNA is applied to one end of a nitrocellulose strip. The DNA migrates to the opposite end of the strip by capillary forces and hybridizes to a specific DNA probe immobilized in a reaction zone (RZ), located in the middle of the strip. Unhybridized DNA migrates away from the RZ. The biotinylated hybrid is visualized by a color reaction employing a streptavidin-alkaline phosphatase (SA-AP) conjugate and a specific chromogenic substrate. The new PACHA technique allows for detection of as little as 1-5 pg of specific human papilloma virus 16 (HPV16) DNA in 25 min of hybridization. In this system, the hybridization efficiency is controlled by the flow velocity of the hybridization solution (HS) and by the volume of the amplified labeled DNA migrating across the immobilized probe. Glycerol (30%) or polyvinyl pyrrolidone (PVP) (1%) reduces the flow rate by a factor of 2.5-3 and increases the sensitivity of the assay by a factor of 5.2 for glycerol and 2.6 for PVP. This novel method ensures efficient hybridization to multiple probes and appears to be superior to currently available solid-phase hybridization techniques.

Base Sequence↗
PubMed1992

Parametric representation of normal breath sounds.

The spectral content of normal tracheal and chest wall breath sounds has been calculated using the fast Fourier transform (FFT) (J. Appl. Physiol. 50: 307-314, 1981). Parameter estimation methods, in particular autoregressive (AR) modeling, are alternative techniques for measuring lung sounds. The outcome of AR modeling of 38 complete breaths picked up simultaneously over the chest walls and tracheae of five normal males was evaluated. The sounds were treated as noise, bounded by a quasi-periodic envelope generated by the cyclic action of breathing, thus causing the sounds to become inherently nonstationary. Normalization of the sounds to their corresponding variance envelopes eliminated the nonstationarity, an important requirement for most signal-processing methods. Subsequently, the AR model order was sought using formal criteria. Orders 6-8 were found to be suitable for normal chest wall sounds, whereas tracheal sounds required at least orders 12-16. Using orders 6 and 12, we compared the prominent spectral features of chest wall and tracheal sounds calculated by AR with those found in the spectra calculated by FFT. The polar representation of the AR roots, calculated from the AR coefficients, showed that normal lung sounds from a group of individuals are characterized by a low variability, suggesting that this method may provide an alternative representation of the sounds. The data presented here show that normal lung sounds, when measured in the frequency domain by either FFT or AR modeling, have a characteristic pattern that is independent of the analysis method.

Adult↗
PubMed1990

Zinc excretion in osteoporotic women.

The relation of zinc to the aging skeleton was investigated in 140 women aged 36-85 years, mostly postmenopausal, who attended the Jerusalem Osteoporosis Center. Osteoporosis was determined by lumbar spine radiograms (Smith index). Bone density (BD) of the distal radius was assessed by Compton spectroscopy and bone mineral content (BMC) at the same site by single-photon absorptiometry. Urine samples (24 h) were analyzed for zinc (UZn), hydroxyproline (UHP), calcium (UCa), magnesium (UMg), and phosphorus (UP) and expressed per gram creatinine. Patients with definite osteoporosis (n = 94) compared to subjects with borderline or no osteoporosis (n = 34) had a significantly higher mean age (67.4 versus 58.6 years), postmenopausal age (PMA, 19.9 versus 11.0), UZn (811 versus 581), UHP (23.5 versus 18.2), and UMg (90.4 versus 74.3). Urinary calcium UCa and phosphorus UP were similar in both groups. The bone mass measurements BD, BMC, and CI were lower in the osteoporotic group. Hyperzincuria (UZn above 800 micrograms/g creatinine) was found in 41 osteoporotic patients (45%) compared to 6 subjects in the control group (17%). In view of the positive correlation between UZn and age (r = 0.35, p = 0.001) and to eliminate the effect of age, a separate analysis was performed for 66 subjects under the age of 65 in whom the mean age was similar for the osteoporosis patients (n = 38) and control group (n = 28). Nevertheless, the osteoporosis patients still had a significantly higher mean UZn and UHP.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1990

Hybricomb. A novel diagnostic tool for DNA probing.

A technique is described in which unlabeled DNA probes are immobilized on the plastic surface of a 12-tooth comb and used to capture homologs by a hybridization procedure. In the examples described, cellular DNA from cervical biopsies is chemically labeled using the Chemi-probe system. The sample containing labeled DNA is then hybridized with the immobilized unlabeled probe (reverse hybridization). By means of this technique, human papillomavirus sequences can be detected with a sensitivity comparable to that of radioactive probe procedures. DNA hybrids are visualized as colored spots on each tooth by moving the comb through the reagent solutions of the prefilled developing plate. The whole procedure requires less than 50 minutes hands-on time, and the results are obtained in a few hours. Application of the technique to the detection and typing of mycoplasma DNA is also reported.

Bacterial Typing Techniques↗
PubMed1989

Chemiprobe, a nonradioactive system for labeling nucleic acid. Principles and applications.

The Chemiprobe Kit provides a complete system for nonradioactive labeling of DNA probes and their detection in hybridization studies. The system is highly sensitive, permitting the detection of 0.2-0.4 pg DNA which allows detection of a single gene sequence in 0.5-1 microgram of bacterial DNA or in 3-5 micrograms of mammalian DNA. In this paper the authors show that the rRNA genes of M. capricolum can be detected by using only 50 ng/ml of sulfonated probe cloned from another mycoplasma, M. pneumoniae. The Chemiprobe system has been successfully used in the detection of the single copy human gene for glucocerobrosidase from total embryonic DNA by hybridization to a specific sulfonated cDNA. 5 x 10(4) M. pneumoniae cells can be detected either free or mixed with sputum using a standard dot blot technique: mycoplasma cells were lysed by a mucolytic agent, denaturated by NaOH, immobilized on a nylon membrane filter, and then hybridized with pPN4, a plasmid DNA probe specific for M. pneumoniae. The resulting hybrids were then detected by the standard Chemiprobe procedure. A new kit based on the Chemiprobe system has been designed especially for the detection of mycoplasmas in tissue culture. This kit has been tested on 70 random samples collected from tissue culture fluids from 11 different sources. Of these, 42 were found to be contaminated by the Chemiprobe procedure, whereas 41 were found to be contaminated by classical microbiological methods. No false negatives were found.

DNA↗
PubMed1987

The detection of neurotoxicant activity by a bacterial toxicity assay.

We present and evaluate a simple, rapid (2 hrs), colorimetric assay for the detection of toxicants, including neurotoxicants. The assay is based on the ability of toxicants to inhibit the de novo synthesis of an inducible enzyme, beta-galactosidase, by a rough mutant of E. coli, which is highly sensitive to a wide spectrum of toxic substances. The test is performed under stress conditions for the bacterium, since under such conditions better sensitivity for low concentrations of analytes is obtained. In the assay, the sample is mixed with the stressed bacteria and a cocktail containing the specific inducer for the chromogenic enzyme and factors essential to the recovery of the bacteria from their stressed condition. The ability of cells to synthesize B-galactosidase under these conditions depends on their ability to recover from the stress. Toxic materials interfere and/or inhibit the recovery process and with it the synthesis of beta-galactosidase.

Animal Testing Alternatives↗
PubMed1986

'Key paper' for covalent binding of proteins and its uses.

A simple method for covalent coupling of proteins to filter paper modified with quinone groups is described. This paper, termed 'Key paper' is flexible, stable on storage and does not require any activation before use. Proteins bound to Key paper can be detected by enzyme immunoassay, radioimmunoassay or Coomassie blue staining. Bound enzymes retain their enzymatic activity. Nucleic acids do not bind and do not interfere with the activity of the bound proteins. Because of its mechanical and chemical properties Key paper is a good matrix for electroblotting and for direct in situ analysis of proteins.

Antibodies↗
PubMed1985

Double stranded RNA and the nuclear matrix--implications for the 2-5A system.

RNA species present on rat liver nuclear matrices were investigated. Nuclear matrices prepared by extensive digestion of isolated nucleii with DNase and RNaseA followed by low and high (2M NaC1) salt washes were labelled in vitro with T4 RNA ligase and [5'-32P]pCp and the labelled RNA analysed by gel electrophoresis. Despite the extensive RNaseA treatment, a prominent RNA species migrating as a heterodisperse band of 220-300 nucleotides (termed MX220-300), was observed--only minor amounts of other RNA molecules were seen. A comparison of RNA isolated from in-vitro labelled nuclear matrices with isolated matrix RNA that was subsequently labelled, indicated that part of MX220-300 was preferentially exposed on the nuclear matrix structure. Analysis of MX220-300 indicated that it was composed of a polyadenylic acid moiety hydrogen bonded to a smaller molecule of polyuridylic acid. No evidence was found for the presence of guanosine or cytosine residues. Control experiments in which labelled polyuridylic acid was added to nucleii prior to the preparation of MX220-300, virtually excluded the possibility that the partial double stranded RNA structure was an artefact of matrix preparation. An analysis of proteins in the nuclear matrix structure that interact with double stranded (ds)RNA showed at least 2 proteins having molecular weights of 62K and 66K daltons that recognized and bound polyadenylic/polyuridylic acid. Competition experiments with unlabelled polyinosinic/polycytidylic acid indicated that these proteins specifically recognized the dsRNA structure. The 62K and 66K dalton matrix proteins that specifically bound dsRNA were observed in nuclear matrices prepared from HeLa, Ehrlich ascites tumor and rat liver cells. It is not known whether these matrix located dsRNA binding proteins have 2-5A synthetase activity. The relevance of the above findings to the 2-5A system will be discussed.

Adenine Nucleotides↗
PubMed1984

A new blotting medium for the simple isolation and identification of highly resolved messenger RNA.

A simple method has been developed that allows the rapid isolation and identification of highly resolved mRNA molecules. RNA species are separated by gel electrophoresis and then blotted on to a paper sheet to which polyuridylic acid has been covalently bound. This mRNA affinity paper ("mAP") specifically binds, in a reversible manner, polyA+ containing molecules. A replica picture of the agarose gel is thus obtained on the mAP, from which bound mRNA molecules can be eluted by heating in water. In addition to their simple isolation individual mRNA species, whilst still bound to mAP, can be identified by both "in-situ" hybridization and translation.

Animals↗
PubMed1984

Isolation of poly(A)+ RNA by paper affinity chromatography.

Poly(A)+ RNA was isolated from in vitro short-term-labeled total cytoplasmic RNA of Ehrlich ascites tumor cells by oligo(dT) cellulose chromatography. This poly(A)+ RNA fraction was compared with a poly(A)+ RNA fraction isolated by a new procedure which involves specific binding of poly(A)+ RNA to messenger affinity paper (mAP) and its release in hot (70 degrees C) water. In typical experiments 10-11 micrograms (2.3%) of poly(A)+ RNA can be retained from 500 micrograms of total cytoplasmic RNA per cm2 of mAP in a quick one-step procedure. The poly(A)+ RNA preparations isolated by the two methods proved to be almost identical with respect to their fraction in total cytoplasmic RNA, specific radioactivities, sucrose gradient profiles, and translation assays. Since the isolation of poly(A)+ RNA by mAP is much less time consuming than that by oligo(dT) column chromatography and since the poly(A)+ RNA can be recovered from mAP in small volumes, which avoids further loss during precipitations, it can be advantageously used for preparative isolation of poly(A)+ RNA.

Animals↗
PubMed1983

Location of RNase activity in nuclear residual structures.

We report here the partial isolation of an RNase activity which copurifies with the nuclear residual structure prepared from both rat liver and Herpes-infected BHK cells. After the successive treatment of purified nuclei with DNase, low salt and high salt, the RNase activity is found both in the high salt soluble supernatant fraction and in the residual nuclear structure. Triton X-100 treatment of this structure solubilizes the RNase activity. From this we conclude that some of the RNase activity associated with the nuclear residual structure may be located in either the phospholipidic or protein moieties that were extracted with Triton X-100. This RNase cuts rRNA non-randomly into characteristic degradation products. Its molecular weight, on a glycerol gradient, was determined to be 25,000.

Animals↗
PubMed1981

Efficient DNA transfection and rapid assay for thymidine kinase activity and viral antigenic determinants.

We describe DEAE dextran-mediated DNA transfection in suspension which routinely gives transient gene expression in 0.1--1% of the transfected cells. We have used normal diploid human skin fibroblasts, monkey BSC cells, and mouse L or 3T6 cells with almost equal efficiency. Gene expression is detected 1--3 days after addition of the DNA. SV40 and polyoma T and V antigen are detected by in situ immunofluorescence and thymidine kinase gene expression is detected by in situ autoradiography. The high efficiency of transfection and the speed of detection together provide a means to study transfecting gene functions that does not rely on selection to obtain stable transformants. It should be possible to screen for the expression of any gene product which can be assayed by in situ immunofluorescence or autoradiographic techniques.

Animals↗