Cell attachment and chemotaxis can utilize the same peptide sequence of fibronectin

Fibronectin and fragments of the parent molecule are involved in cell-substrate attachment, cell migration and chemotaxis. The cell attachment sequence, Gly-Arg-Gly-Asp-Ser-Pro, was synthesized and tested for its ability to effect ligamentum nuchae fibroblast chemotaxis. This hexapeptide and fibronectin itself both caused directed cell migration, with maximal activity in the 10(-10) to 10(-9) M range. These data demonstrate that the fibronectin cell binding site and chemotaxis site share a common sequence.     

Identification of a tumor cell receptor for VGVAPG, an elastin-derived chemotactic peptide

Extracellular matrix proteins and their proteolytic products have been shown to modulate cell motility. We have found that certain tumor cells display a chemotactic response to degradation products of the matrix protein elastin, and to an elastin-derived peptide, VGVAPG. The hexapeptide VGVAPG is a particularly potent chemotaxin for lung-colonizing Lewis lung carcinoma cells (line M27), with 5 nM VGVAPG eliciting maximal chemotactic response when assayed in 48-microwell chemotaxis chambers. Binding of the elastin-derived peptide to M27 cells was studied using a tyrosinated analog (Y-VGVAPG) to allow iodination. Scatchard analysis of [125I]Y-VGVAPG binding to viable M27 tumor cells at both 37 and 4 degrees C indicates the presence of a single class of high affinity binding sites. The dissociation constant obtained from these studies (2.7 X 10(-9) M) is equivalent to the concentration of VGVAPG required for chemotactic activity. The receptor molecule was identified as an Mr 59,000 species by covalent cross-linking of the radiolabeled ligand to the M27 tumor cell surface and subsequent analysis of the cross-linked material by electrophoresis and size-exclusion high performance liquid chromatography. These results suggest that M27 tumor cell chemotaxis to VGVAPG is initiated by high affinity binding of the peptide to a distinct cell surface receptor.     

The dog common carotid artery: a sensitive bioassay for studying vasodilator effects of substance P and of kinins

In order to develop a sensitive pharmacological preparation which would allow the measurement of the inhibitory effects of kinins and substance P (SP) in vascular smooth muscles, several large arteries of the dog were studied in vitro. The common carotid artery was found to be one of the most sensitive preparations to SP and kinins. When contracted with low concentrations of noradrenaline (between 3.0 x 10(-8) and 3.0 x 10(-7) M), this artery responds to SP (6.5 x 10(-11)-6.5 x 10(-9) M) and bradykinin (BK) (8.1 x 10(-11)-9.1 x 10(-8) M) with relaxations that are proportional to the concentrations of the two peptides. SP and BK appear to exert their relaxant effects through the activation of specific receptors as the exposure of the common carotid artery to concentrations of [Leu8]-angiotensin II, propranolol, methysergide, cimetidine, or atropine sufficient to inhibit the effects of the corresponding agonists do not affect the relaxing effect of SP and BK. [Leu8]-des-Arg9-BK (1.0 x 10(-6) M), indomethacin (2.8 x 10(-5) M), and lioresal (4.7 x 10(-5) M) are also inactive. When the dog common carotid artery is desensitized with high concentrations of SP, BK, eledoisin, and physalaemin a cross-desensitization is observed only between SP and physalaemin. These results support the conclusion that SP and kinins act on different receptors. The order of potency of kinins is the following: BK = [Tyr(Me)8]-BK greater than des-Arg9-BK, suggesting that the receptor for kinins is of the B2 type. The order of potency of peptides related to SP is SP greater than C-terminal 4-11 greater than C-terminal hexapeptide 6-11, similar to that observed in other vascular preparations. The results summarized in this paper indicate that the dog common carotid artery is a preparation sensitive to SP and BK and useful for studying the relaxant effect of these two peptides on vascular smooth muscles.     

The inhibitory effect of hydrocortisone on interferon production by rat spleen cells

The effect of hydrocortisone on interferon r(IFN-r) production by rat spleen cells and its mechanism were studied. The results showed that hydrocortisone inhibited IFN-r production at concentrations as low as 5.52 x 10(-10) M, with complete suppression at 5.52 x 10(-8) M, and the total number and survival rate of the cultured spleen cells were not apparently affected by 5.52 x 10(-8) M hydrocortisone. The inhibitory effect was dose-dependent when the concentration was from 5.52 x 10(-10) M to 5.52 x 10(-8) M and could be blocked by RU38486, a competitive antagonist of glucocorticoid. Our results suggested that glucocorticoid may inhibit IFN-r production through a receptor-mediated mechanism.     

Thrombin-induced chemotaxis and aggregation of neutrophils

Thrombin-induced neutrophil chemotaxis and aggregation were studied using cells isolated from either human or sheep blood. Sheep neutrophils (10(8) cells/ml) exhibited maximum chemotactic migration towards 10(-8)M human alpha-thrombin, 10(-8)M gamma-thrombin (which lacks the fibrinogen site), and 10(-12)MD-Phe-Pro-Arg-CH2-alpha-thrombin (catalytically inactive thrombin). Chemotactic responses of the same magnitude were obtained with human neutrophils (10(8) cells/ml). The chemotactic responses to thrombin were comparable to those obtained with diluted (1:200 v/v) zymosan activated serum (ZAS) and 10(-11)M FMLP. Premixing of the thrombin forms with hirudin in 1:1 stoichiometric amounts abolished the chemotaxis but not chemokinesis Aggregatory responses of human and sheep neutrophils were comparable for ZAS, alpha-thrombin, and gamma-thrombin. The responses of both human and sheep neutrophils to D-Phe-Pro-Arg-CH2-alpha-thrombin were attenuated, indicating that the proteolytic site may be involved in the aggregatory response. The results suggest that thrombin-induced neutrophil chemotaxis and aggregation are mediated by different mechanisms, since chemotaxis is a catalytically independent response whereas aggregation is an active site independent response.     

The N-terminal hexapeptide fragment of IGF II stimulates thymidine incorporation into fibroblasts.

We synthesized the N-terminal hexapeptide fragment of IGF II to study potential binding to NMDA receptors in analogy to the N-terminal tripeptide of IGF I. The amino acid sequence of the hexapeptide is furthermore identical with the C-terminal sequence of the casiragua insulin B chain. The hexapeptide did not bind to the NMDA receptors, but was found to promote [3H]-thymidine incorporation into fibroblasts at concentrations of 10(-8) - 10(-5) M in a dose-dependent manner. Since [125I]-hexapeptide did not bind to IGF receptors, indirect competition studies using either labelled IGFs or insulin had to be used. The competition of hexapeptide at a concentration of 10(-5) M with labelled IGF I or II was about equal to that of 10(-9) M IGF I or II. IGF receptors were apparently up-regulated by the hexapeptide, as has also been described for insulin. When using casiragua insulin as labelled ligand, IGF II and casiragua insulin competed with equal potency, whereas the hexapeptide at 10(-7) M caused an apparent up-regulation of the casiragua insulin binding sites. Our results that the hexapeptide stimulates [3H]-thymidine incorporation and up-regulates IGF II and casiragua insulin binding sites may be connected to one or several of the following findings: the hystricomorph insulins--of which the casiragua insulin is a member--stimulate DNA synthesis to a greater extent than other insulins; the insulin and type 1 IGF receptor binding regions are localized predominantly in the C-terminal region of the insulin B chain; and the "cooperative" site regulating the affinity of the insulin receptor is also located in the C-terminal region of the insulin B chain. Further experiments will be needed to clarify the exact mechanism.     

Release of prostaglandin E2 and thromboxane B2 by mixed isolated human lung cells

Human lung specimens were minced and treated for 30 min with collagenase (1 mg ml-1) and DNase (0.1 mg ml-1) to obtain a suspension of viable (approximately 80%) and metabolically active lung cells (5 x 10(6) cells per gram of tissue). Treatment of these mixed lung cells with bradykinin (1.25 x 10(-6) to 1 x 10(-5) M) and f-Met-Leu-Phe (f-MLP; 1 x 10(-8) to 5 x 10(-6) M) did not stimulate to a substantial extent the release of prostaglandins and thromboxanes (measured with novel Enzyme Immunoassays). The only concentration of PAF that stimulated significantly the release of icosanoids from lung cells was 5 x 10(-7) M. Phorbol myristate (PMA; 5 x 10(-8) to 2 x 10(-6) M) and ionophore a-21387 (2.5 x 10(-6) to 2 x 10(-5) M) strongly stimulated the release of prostaglandins and thromboxanes by dispersed human lung cells. These findings support previous observations showing that human lungs have the enzymes necessary for the synthesis and release of prostaglandins and thromboxanes but stimulation of the release of these mediators is not obtained with the hormonal stimuli that are active in guinea pigs. Studies in progress will purify the cell populations and characterize the cells responsible for the release of these icosanoids.     

Effects of organic antagonists of Ca(2+), Na(+), and K(+) on chemotaxis and motility of escherichia coli

Various Ca(2+) antagonists used in animal research, many of them known to be Ca(2+) channel blockers, inhibited Escherichia coli chemotaxis (measured as entry of cells into a capillary containing attractant). The most effective of these, acting in the nanomolar range, was omega-conotoxin GVIA. The next most effective were gallopamil and verapamil. At concentrations around 100-fold higher than that needed for inhibition of chemotaxis, each of these antagonists inhibited motility (measured as entry of cells into a capillary lacking attractant). Various other Ca(2+) antagonists were less effective, though chemotaxis was almost always more sensitive to inhibition than was motility. Cells treated with each of these Ca(2+) antagonists swam with a running bias, i.e., tumbling was inhibited. Similarly, some Na(+) antagonists used in animal research inhibited bacterial chemotaxis. E. coli chemotaxis was inhibited by saxitoxin at concentrations above 10(-7) M, while more than 10(-4) M was needed to inhibit motility. Cells treated with saxitoxin swam with a tumbling bias. In the case of other Na(+) antagonists in animals, aconitine inhibited bacterial chemotaxis 10 times more effectively than it inhibited motility, and two others inhibited chemotaxis and motility at about the same concentration. In the case of K(+) antagonists used in animal research, 4-aminopyridine blocked E. coli chemotaxis between 10(-3) M and, totally, 10(-2) M, while motility was not affected at 10(-2) M; on the other hand, tetraethylammonium chloride failed to inhibit either chemotaxis or motility at 10(-2) M.     

Quantitative relationships between single-cell and cell-population model parameters for chemosensory migration responses of alveolar macrophages to C5a

Phenomenological parameters from a mathematical model of cell motility are used to quantitatively characterize chemosensory migration responses of rat alveolar macrophages migrating to C5a in the linear under-agarose assay, simultaneously at the levels of both single cells and cell populations. This model provides theoretical relationships between single-cell and cell-population motility parameters. Our experiments offer a critical test of these theoretical linking relationships, by comparison of results obtained at the cell population level to results obtained at the single-cell level. Random motility of a cell population is characterized by the random motility coefficient, mu (analogous to a particle diffusion coefficient), whereas single-cell random motility is described by cell speed, s, and persistence time, P (related to the period of time that a cell moves in one direction before changing direction). Population chemotaxis is quantified by the chemotactic sensitivity, chi 0, which provides a measure of the minimum attractant gradient necessary to elicit a specified chemotactic response. Single-cell chemotaxis is characterized by the chemotactic index, CI, which ranges from 0 for purely random motility to 1 for perfectly directed motility. Measurements of cell number versus migration distance were analyzed in conjunction with the phenomenological model to determine the population parameters while paths of individual cells in the same experiment were analyzed in order to determine the single-cell parameters. The parameter mu shows a biphasic dependence on C5a concentration with a maximum of 1.9 x 10(-8) cm2/sec at 10(-11) M C5a and relative minima of 0.86 x 10(-8) cm2/sec at 10(-7) M C5a and 1.1 x 10(-8) cm2/sec in the absence of Ca; s and P remain fairly constant with C5a concentration, with s ranging from 2.1 to 2.5 microns/min and P varying from 22 to 32 min. chi 0 is equal to 1.0 x 10(-6) cm/receptor for all C5a concentrations tested, corresponding to 60% correct orientation for a difference of 500 bound C5a receptors across a 20 microns cell length. The maximum CI measured was 0.2. Values for the population parameters mu and chi 0 were calculated from single-cell parameter values using the aforementioned theoretical linking relationships. The values of mu and chi 0 calculated from single-cell parameters agreed with values of mu and chi 0 determined independently from population migrations, over the full range of C5a concentrations, confirming the validity of the linking equations. Experimental confirmation of such relationships between single-cell and cell-population parameters has not previously been reported.     

Effect of procaine hydrochloride on the electrophoretic mobility of human red blood cells

The interaction of cationic anesthetics with biological membranes and the resulting alterations of membrane electrokinetic properties continue to be of current interest. The present study was designed to examine the effects of procaine hydrochloride (PRHCL) on the mobility of human red blood cells (RBC); electrophoretic measurements were made on RBC suspended in phosphate-buffered saline (PBS, pH = 5.0, 7.4, or 9.2), autologous plasma or 3 g% dextran T70/PBS (pH = 7.4), with PRHCL concentrations from 8 x 10(-6) to 8 x 10(-2) M. Low concentrations of PRHCL (8 x 10(-5)-8 x 10(-3) M) significantly (p less than 0.001) increased RBC mobility, with a maximal increase of 8.2% at 8 x 10(-4) M. Conversely, a higher PRHCL concentration (8 x 10(-2) M significantly (p less than 0.001) decreased RBC mobility. Both glutaraldehyde fixation and lipid extraction abolished any PRHCL-induced increase in RBC mobility; the observed increases in mobility for normal cells are, thus, consistent with a mechanism based on expansion of the RBC membrane glycocalyx. Microelectrophoretic methods were also used to study the effect of PRHCL (8 x 10(-4) and 8 x 10(-2) M) on RBC membrane calcium binding, with the results indicating that PRHCL competes with calcium for neuraminate binding sites. We conclude that the observed changes in RBC electrokinetic properties reflect incorporation of PRHCL into the RBC membrane; such changes may be of importance in modulating cell-cell interactions.     

Flow cytometric analysis of bromodeoxyuridine-substituted cells stained with 33258 Hoechst.

This paper describes a flow-cytometric application of the quenching of fluorescence from 33258 Hoechst stained Chinese hamster ovary-line cells due to the incorporation of 5-bromo-deoxyuridine (BrdU) into the cellular deoxyribonucleic acid. Cells were grown for 24 hr in medium containing BrdU in concentrations ranging from 1 x 10(-8) to 1 x 10(-4) M. For each concentration we measured the average fluorescence as determined by flow cytometry, the extent of BrdU substitution and the effect of the BrdU on cell growth. We determined that a BrdU concentration of 1 x 10(-5) M resulted in sufficient substitution to quench the fluorescence from 33258 Hoechst by a factor of 4, allowing discrimination between cycling and noncycling cells. The extent of BrdU substitution after growth for 24 hr in this concentration of BrdU was 64%. These data indicate the feasibility of detecting deoxyribonucleic acid synthesis in whole cells using the 33258 Hoechst-BrdU methodology.     

Amplification effect and mechanism of action of ET-1 in U-46619-induced vasoconstriction in pig skin

The aim of this study was to investigate if a low concentration of endothelin-1 (ET-1; 8 x 10(-10) M) may amplify the skin vasoconstrictor effect of other vasoactive substances in the pathogenesis of skin vasospasm. Pig skin flaps (6 x 16 cm) were perfused with Krebs buffer equilibrated with 95% O(2) and 5% CO(2) at 37 degrees C and pH 7.4. Skin perfusion pressure measured by a pressure transducer and skin perfusion assessed by the dermofluorometry technique were used for assessment of skin vasoconstriction. We observed that ET-1 (8 x 10(-10) M) significantly amplified the concentration-dependent (10(-7)-10(-5) M) skin vasoconstrictor effect of norepinephrine. More importantly, we observed for the first time that this low concentration of ET-1 also amplified the concentration-dependent (10(-8)-10(-6) M) skin vasoconstrictor effect of the thromboxane A(2) mimetic U-46619, and this amplification effect of ET-1 was completely blocked by the protein kinase C (PKC) inhibitor chelerythrine (5 x 10(-6) M). Conversely, the PKC activator phorbol 12,13-dibutyrate (10(-7) M) amplified the vasoconstrictor effect of U-46619. Furthermore, the sensitivity of the skin vasculature to the vasoconstrictor effect of extracellular Ca(2+) in U-46619-induced skin vasoconstriction was significantly enhanced in the presence of 8 x 10(-10) M ET-1. Finally, the cyclooxygenase inhibitor indomethacin (5 x 10(-6) M) did not affect the amplification effect of ET-1 on U-46619-induced skin vasoconstriction. We conclude that a low concentration of ET-1 can amplify the skin vasoconstrictor effect of U-46619 independent of endogenous cyclooxygenase products, and the mechanism may involve activation of PKC and increase in sensitivity of the contractile apparatus to Ca(2+) in smooth muscle cells.     

Prostaglandin E(2) inhibits fibroblast chemotaxis

Fibroblasts are the major source of extracellular connective tissue matrix, and the recruitment, accumulation, and stimulation of these cells are thought to play important roles in both normal healing and the development of fibrosis. Prostaglandin E(2) (PGE(2)) can inhibit this process by blocking fibroblast proliferation and collagen production. The aim of this study was to investigate the inhibitory effect of PGE(2) on human plasma fibronectin (hFN)- and bovine bronchial epithelial cell-conditioned medium (BBEC-CM)-induced chemotaxis of human fetal lung fibroblasts (HFL1). Using the Boyden blind well chamber technique, PGE(2) (10(-7) M) inhibited chemotaxis to hFN 40.8 +/- 5.3% (P < 0.05) and to BBEC-CM 49.7 +/- 11.7% (P < 0.05). Checkerboard analysis demonstrated inhibition of both chemotaxis and chemokinesis. The effect of PGE(2) was concentration dependent, and the inhibitory effect diminished with time. Other agents that increased fibroblast cAMP levels, including isoproterenol (10(-5) M), dibutyryl cAMP (10(-5) M), and forskolin (3 x 10(-5) M) had similar effects and inhibited chemotaxis 54.1, 95.3, and 87.0%, respectively. The inhibitory effect of PGE(2) on HFL1 cell chemotaxis was inhibited by the cAMP-dependent protein kinase (PKA) inhibitor KT-5720, which suggests a cAMP-dependent effect mediated by PKA. In summary, PGE(2) appears to inhibit fibroblast chemotaxis, perhaps by modulating the rate of fibroblast migration. Such an effect may contribute to regulation of the wound healing response after injury.     

Comparative study of oestrogenic properties of eight phytoestrogens in MCF7 human breast cancer cells

Previous studies have compared the oestrogenic properties of phytoestrogens in a wide variety of disparate assays. Since not all phytoestrogens have been tested in each assay, this makes inter-study comparisons and ranking oestrogenic potency difficult. In this report, we have compared the oestrogen agonist and antagonist activity of eight phytoestrogens (genistein, daidzein, equol, miroestrol, deoxymiroestrol, 8-prenylnaringenin, coumestrol and resveratrol) in a range of assays all based within the same receptor and cellular context of the MCF7 human breast cancer cell line. The relative binding of each phytoestrogen to oestrogen receptor (ER) of MCF7 cytosol was calculated from the molar excess needed for 50% inhibition of 3H]oestradiol binding (IC50), and was in the order coumestrol (35x)/8-prenylnaringenin (45x)/deoxymiroestrol (50x)>miroestrol (260x)>genistein (1000x)>equol (4000x)>daidzein (not achieved: 40% inhibition at 10(4)-fold molar excess)>resveratrol (not achieved: 10% inhibition at 10(5)-fold molar excess). For cell-based assays, the rank order of potency (estimated in terms of the concentration needed to achieve a response equivalent to 50% of that found with 17beta-oestradiol (IC50)) remained very similar for all the assays whether measuring ligand ability to induce a stably transfected oestrogen-responsive ERE-CAT reporter gene, cell growth in terms of proliferation rate after 7 days or cell growth in terms of saturation density after 14 days. The IC50 values for these three assays in order were for 17beta-oestradiol (1 x 10(-11)M, 1 x 10(-11)M, 2 x 10(-11)M), and in rank order of potency for the phytoestrogens, deoxymiroestrol (1 x 10(-10)M, 3 x 10(-11)M, 2 x 10(-11)M)>miroestrol (3 x 10(-10)M, 2 x 10(-10)M, 8 x 10(-11)M)>8-prenylnaringenin (1 x 10(-9)M, 3 x 10(-10)M, 3 x 10(-10)M)>coumestrol (3 x 10(-8)M, 2 x 10(-8)M, 3 x 10(-8)M)>genistein (4 x 10(-8)M, 2 x 10(-8)M, 1 x 10(-8)M)/equol (1 x 10(-7)M, 3 x 10(-8)M, 2 x 10(-8)M)>daidzein (3 x 10(-7)M, 2 x 10(-7)M, 4 x 10(-8)M)>resveratrol (4 x 10(-6)M, not achieved, not achieved). Despite using the same receptor context of the MCF7 cells, this rank order differed from that determined from receptor binding. The most marked difference was for coumestrol and 8-prenylnaringenin which both displayed a relatively potent ability to displace [3H]oestradiol from cytosolic ER compared with their much lower activity in the cell-based assays. Albeit at varying concentrations, seven of the eight phytoestrogens (all except resveratrol) gave similar maximal responses to that given by 17beta-oestradiol in cell-based assays which makes them full oestrogen agonists. We found no evidence for any oestrogen antagonist action of any of these phytoestrogens at concentrations of up to 10(-6)M on either reporter gene induction or on stimulation of cell growth.     

Differential effect of DHEA on mitogen-induced proliferation of T and B lymphocytes

Dehydroepiandrosterone (DHEA) is the predominant steroid hormone secreted by adrenal gland, and it has been proposed in recent years that DHEA has significant effects on immune function. We investigated the effect of DHEA (1 x 10(-5) - 1 x 10(-8)M) on proliferation of human T cells and B cells and on immunoglobulin production, a representative function of B cells. High doses of DHEA (1 x 10(-5)) significantly inhibited proliferation of peripheral blood mononuclear cells (PBMCs) and T cells induced by T cell mitogens hemagglutinin (PHA) and concanavalin A (Con A). Proliferation of PBMCs induced by B cell mitogens pokeweed mitogen (PWM) was increased by 1 x 10(-7) - 1 x 10(-6)M DHEA. Proliferation of PBMCs and B cells induced by Staphylococcus aureus Cowan strain I (SAC) was not significantly changed at any concentrations of DHEA. However, a concentration of 1 x 10(-7)M DHEA tended to potentiate their proliferation. This study suggested that DHEA acted on T and B lymphocytes differentially in immune system.     

Antimutagenic action of cobaltous chloride on radiation-induced mutations in cultured Chinese hamster cells

The effects of cobaltous chloride on 8-azaguanine (8AG)-resistant mutations induced by gamma-rays or ultraviolet (UV) light in cultured Chinese hamster V79 cells were examined. Cobaltous chloride alone had no significant effects on survival and mutations of V79 cells at concentrations less than 1 x 10(-5) M. Cobaltous chloride at a concentration of 3 x 10(-6) M had a marked effect in reducing 8AG-resistant mutations induced by gamma-rays of 2-6 Gy, when cells were incubated for 6-7 days in the presence of cobaltous chloride after gamma-ray irradiation (posttreatment). The pretreatment of cells with cobaltous chloride for 6 days before gamma-ray irradiation reduced 8AG-resistant mutations induced by gamma-rays. Pre- or post-treatment with cobaltous chloride had no such effect on UV-induced mutations, however. The difference in responsiveness to cobaltous chloride between bacterial and mammalian cell systems is discussed.     

The antimitotic effect of the neem terpenoid azadirachtin on cultured insect cells

When cultured insect cells (Sf9) were grown in the presence of 5 x 10(-6) M azadirachtin, there was a rapid increase in the mitotic index, with the appearance of many aberrant mitotic figures. Flow cytometry established that cells accumulated in the G2/M phase of the cell cycle, and that the effect was concentration-dependent. At 10(-8) M a period of 20 h was necessary to raise the proportion in G2/M to 42% above the control values, but at 5 x 10(-6) M more than 90% of the cells were in this phase. Azadirachtin had the same effect on C6/36 mosquito cells, but failed to affect L929 murine fibroblast cells even at a concentration of 10(-4) M over 72 h. Experiments with colchcine and taxol showed similarities of action between azadirachtin and colchicine, and azadirachtin was apparently able to displace colchicine-fluorescein from binding-sites in living insect cells. Another similarity between azdirachtin and colchicine was that both phytochemicals prevented the polymerisatrion in vitro of mammalian tubulin, although the azadirachtin was much less effective.     

Differences among primates in defence against infection: sensitivity of polymorphonuclear leukocytes to fMet-Leu-Phe

The sensitivity of polymorphonuclear leukocytes (PMN) to N-formyl-methionyl-leucyl-phenylalanine (fMet-Leu-Phe) for chemotaxis and for lysosomal enzyme release was examined using the PMN of four primate species, human (H. sapiens), chimpanzee (P. troglodytes), rhesus monkey (M. mulatta), and cotton-headed tamarin (S. (O) oedipus). The 50 per cent effective concentrations (EC50) of fMet-Leu-Phe for chemotaxis were 2.5 X 10(-9) M in human, 10(-9) M in chimpanzee, 8 X 10(-8) M in rhesus monkey, and 3.3 X 10(-6) M in tamarin. The EC50 values of fMet-Leu-Phe for myeloperoxidase (MPO) release were 10(-8) M in human, 4 X 10(-8) M in chimpanzee, 4 X 10(-8) M in rhesus monkey, and 10(-6) M in tamarin and those for beta-glucuronidase release were 4 X 10(-9) M, 6.4 X 10(-8) M, 1.8 X 10(-7) M, and 1.6 X 10(-6) M, respectively. Thus, the sensitivity to fMet-Leu-Phe for chemotaxis was in the order: chimpanzee congruent to human greater than rhesus monkey greater than tamarin, and that for the release of lysosomal enzymes, MPO and beta-glucuronidase, was in the order: human greater than chimpanzee greater than rhesus monkey greater than tamarin. These results appear to indicate that the sensitivity to fMet-Leu-Phe increases in the order of evolution of primates toward the human, and suggest that the sensitivity of PMN in the defence function against infection also increases in the same order.     

Anti-thyroid hormonal activity of tetrabromobisphenol A, a flame retardant, and related compounds: Affinity to the mammalian thyroid hormone receptor, and effect on tadpole metamorphosis

The thyroid hormone-disrupting activity of tetrabromobisphenol A (TBBPA), a flame retardant, and related compounds was examined. TBBPA, tetrachlorobisphenol A (TCBPA), tetramethylbisphenol A (TMBPA) and 3,3'-dimethylbisphenol A (DMBPA) markedly inhibited the binding of triiodothyronine (T3; 1 x 10(-10) M) to thyroid hormone receptor in the concentration range of 1 x 10(-7)-1 x 10(-4) M, while bisphenol A and 2,2-diphenylpropane were inactive. TBBPA, TCBPA, TMBPA and DMBPA did not exhibit thyroid hormonal activity in a thyroid hormone-responsive reporter assay using a Chinese hamster ovary cell line (CHO-K1) transfected with thyroid hormone receptor alpha1 or beta1, but TBBPA and TCBPA showed significant anti-thyroid hormone effects on the activity of T3 (1 x 10(-8) M) in the concentration range of 3 x 10(-6) - 5 x 10(-5) M. The thyroid hormone-disrupting activity of TBBPA was also examined in terms of the effect on amphibian metamorphosis stimulated by thyroid hormone. TBBPA in the concentration range of 1 x 10(-8) to 1 x 10(-6) M showed suppressive action on T3 (5 x 10(-8) M)-enhancement of Rana rugosa tadpole tail shortening. These facts suggest that TBBPA, TCBPA, TMBPA and DMBPA can act as thyroid hormone-disrupting agents.