Sympathectomy enhances the substance P-mediated breakdown of the blood-aqueous barrier in response to infrared irradiation of the rabbit iris

Summary Rabbits were subjected to infrared irradiation of the iris 1 month after unilateral cervical sympathectomy. The resulting breakdown of the blood-aqueous barrier was greatly enhanced on the sympathectomized side. In contrast, the response to intravitreally injected substance P (SP) was the same in both eyes. The enhancement of the response to IR irradiation could be abolished by pretreatment with an SP antagonist, (D-Pro², D-Trp^7,9)-SP.This study was supported by grants from the Swedish MRC (04X-1007, 14X-2321), the Medical Faculty of Lund, Ferring Arzneimitel, Kiel (Federal Republic of Germany) and Ferring Pharmaceuticals, Malmö (Sweden).     

Development of a new tachykinin antagonist

Summary The mouse urinary bladder possesses a tachykinin receptor which responds to kassinin and eledoisin, but not to some other tachykinins. The action of kassinin, but not that of eledoisin, was blocked in a surmountable manner by a new tachykinin antagonist, D-Pro⁴, Val⁸, D-Trp^7,9,10 (SP)^4–11.Acknowledgments. We thank Mrs Danielle Solomos for technical help. This study was financed by a grant from the Swiss National Science Foundation (3.800-80.2).     

Sympathectomy enhances the substance P-mediated breakdown of the blood-aqueous barrier in response to infrared irradiation of the rabbit iris

Rabbits were subjected to infrared irradiation of the iris 1 month after unilateral cervical sympathectomy. The resulting breakdown of the blood-aqueous barrier was greatly enhanced on the sympathectomized side. In contrast, the response to intravitreally injected substance P (SP) was the same in both eyes. The enhancement of the response to IR irradiation could be abolished by pretreatment with an SP antagonist, (D-Pro2, D-Trp7,9)-SP.     

Influence of oxygen concentration on development ofDrosophila melanogaster

Summary 1st, 2nd, and early 3rd instarDrosophila larvae are extremely sensitive to 100% O₂ or 75% O₂/25% N₂ (at atmospheric pressure) whereas eggs, late 3rd instar larvae, and pupae are relatively insensitive under our exposure conditions. Eclosing flies exposed to an O₂ enriched environment consistently possessed 2 eye abnormalities: dark eye color and altered eye shape.Supported by NSF-URP grant SP1782684 and a Summer Faculty Fellowship to Harry Nickla.     

Effect of an LH-RH antagonist on reproductive status of immature female rats

Summary Administration of the LH-RH antagonist, D-Phe²-D-Ala⁶-LH-RH (Wy-18, 185) to immature female rats from days 25–35 of age was without significant effect on day of vaginal opening (puberal onset), weights of the ovaries, uteri and anterior pituitary, and on ovarian histology on autopsy day 39.The technical expertise of J. Tracy is gratefully acknowleged.     

A glucagon-secretin-like peptide stimulates the intrinsic nervous plexus of guinea pig gallbladder

Summary The role of vasoactive intestinal peptide (VIP), as a possible neurotransmitter of the intrinsic nerve plexus in the guinea pig gallbladder, was investigated by monitoring spontaneous contractile activity. VIP receptor antagonist (4 Cl-D-Phe⁶, Leu¹⁷)-VIP did not produce any effect on muscular tone and spontaneous activity, whereas (N-Ac-Tyr¹, D-Phe²)-GRF-(1-29)-NH₂, (14-GRF analog), which is known to stimulate digestive enzyme secretion by interacting with the VIP-preferring receptors, greatly increased the amplitude and frequency of waves as well as the muscular tone. Since VIP receptor antagonist acts selectively as a competitive antagonist for the action of VIP, we conclude that the gallbladder inhibitory intrinsic plexus neurotransmitter is not VIP, but a member of the glucagon-secretin family of peptides.     

Synthesis and contractile activity of the C-terminal heptapeptide of substance P with N5-dimethyl glutamine in the 6-position. Active site studies

Summary The synthesis and testing of [N⁵-dimethyl-Gln⁶]-SP_5–11 showed 37±12% contractile activity relative to SP, and intrinsic efficacy 98±4%. This finding indicates that the carboxamide groups of the dual Gln⁵-Gln⁶ moiety are not equally related with the contractile response of the C-terminal heptapeptide of SP.The authors wish to express their deep appreciation to Professor H. Niedrich and Dr J. Bergmann of the Institute of Drug Research, Academy of Sciences of DDR, Berlin, for their valuable help in providing the biological data.     

Differential chemotactic potential of mouse platelet basic protein for thymocyte subsets

Mouse platelet basic protein (CXCL7/mPBP) was cloned from thymic stromal cells and further identification indicated that it was expressed in thymic monocytes/macrophages (Mo/Ms). Recombinant mPBP was chemoattractive for target cells of polymorphonuclear leucocytes, peritoneal Mo/Ms and splenic lymphocytes with distinct potencies. CXCR2 was identified to be a cognate receptor for mPBP. Mouse thymocyte subsets of CD4^-CD8^- double-negative (DN), CD4⁺CD8⁺ double-positive (DP), CD4⁺CD8^- single-positive (CD4SP) and CD4^-CD8⁺ single-positive (CD8SP) expressed cell surface CXCR2 with different positive percentages and expression levels. mPBP was chemoattractive for thymocyte subsets with the potency order DN>DP> CD8SP>CD4SP, consistent with the levels of CXCR2 expressed on the respective cells. Thus, mPBP in thymus is functionally redundant with chemokine CXCL12/ SDF-1. Moreover, our finding that thymic Mo/Ms can produce mPBP implies that they may have other functions apart from acting as scavengers in thymus.Received 25 March 2004; received after revision 10 May 2004; accepted 8 June 2004     

Selective effect of burn injury on splenic CD11c+ dendritic cells and CD8α+CD4−CD11c+ dendritic cell subsets

Burn injury causes an immunosuppression associated with suppressed adaptive immune function. Dendritic cells (DCs) are APCs for which signaling via their Toll-like receptors (TLRs) induces their maturation and activation, which is essential for the adaptive immune response. In this study, we examined if burn injury alters the TLR activity of splenic DCs. After injury, we noticed that DC functions were impaired, characterized by a suppressed capacity to prime naive T cells when triggering the TLR4 signaling cascade using specific ligands (LPS or rHSP60). The observed perturbations on LPS-primed DCs isolated from burned mice exhibited significantly diminished IL-12p40 production and enhanced IL-10 secretion-associated impairment in mitogen-activated protein kinase activation. Interestingly, we observed a decrease of TLR4/MD-2 expression on the CD8α⁺ DC subset that persisted following LPS stimulation. The altered TLR4 expression on LPS-stimulated CD8α⁺ DCs was associated with reduced capacity to produce IL-12 after stimulation. Our results suggested that TLR4 reactivity on DCs, especially CD8α⁺ DCs, is disturbed after burn injury.     

The action of the cholecystokinin-A receptor antagonist,devazepide, on the digestive system of the chicken

The influence of the cholecystokinin (CCK)-A receptor antagonist, devazepide (DVZ), on the chicken digestive tract was investigated. The passage of food from the crops of birds treated with DVZ was not significantly different from that of the control. DVZ treatment did not inhibit the biliary flow stimulated by the CCK analogue, caerulein. Dispersed chicken pancreatic acini stimulated with CCK were treated with various concentrations of DVZ. At 10^–5 M, DVZ completely inhibited amylase release; this concentration was much higher than those reported to have similar effects in mammals. The results suggest that the action DVZ as a CCK antagonist in the chicken is very weak.     

Quercetin interacts with calmodulin,a calcium regulatory protein

Summary Quercetin was found to interact with the Ca²⁺-calmodulin complex, suggesting that it may act as a calmodulin antagonist in vivo to inhibit calmodulin-dependent phenomena caused by biologically active agents, including tumor promoters.This work was supported in part by grants-in-aid for scientific research from the Ministry of Education, Science and Culture, Japan.     

EP2 receptor stimulation promotes calcium responses in astrocytes via activation of the adenylyl cyclase pathway

Astrocytes are a heterogeneous population of cells that are endowed with a great variety of receptors for neurotransmitters and neuromodulators. Recently prostaglandin E₂ has attracted great interest since it is not only released by astrocytes but also activates receptors coupled to either phospholipase C or adenylyl cyclase. We report that EP₂ receptor stimulation triggers cAMP production but also causes release of Ca²⁺ from intracellular stores. This effect is shared by other receptors similarly coupled to adenylyl cyclase and elicited by direct stimulation of the enzyme or application of cAMP analogues. However, the stimulation of the Ca²⁺ response by cAMP is not mediated by protein kinase A, since a specific antagonist of this kinase had no effect. Such a cross-talk between cAMP and Ca²⁺ was not observed in all astrocytes. It might therefore reflect a specific resource of either a subpopulation or astrocytes in a specific functional state. Received 6 June 2006; received after revision 25 July 2006; accepted 31 August 2006     

Ocular involvement in hamsters transplanted with a human leukemic T-cell line

Summary A leukemic T-cell line (TALL-1) was serially transplanted for 5 passages into newborn hamsters treated with antilymphocyte serum. This cell line was derived from a leukemic patient with clinical evidence of ocular involvement. I.p. implantation of 1–3×10⁷ cells resulted in disseminated growth of tumors in all 15 recipients after 23–41 days and 8 of them showed leukemic infiltration of the uveal tract of one or both eyes.Supported by a Cancer Research Grant from the Ministry of Health and Welfare of Japan.     

Protective effect of Shigyaku-to,a traditional Chinese herbal medicine,on the infection of herpes simplex virus type 1 (HSV-1) in mice

The antiviral activity of Shigyaku-to (TJS-109), a traditional Chinese herbal medicine, was investigated in mice infected with herpes simplex virus type 1 (HSV-1). TJS-109 is a combination of the medicinal plant extracts fromZingiberis siccatum rhizoma,Aconiti tuber andGlycyrrhizae radix in a specific proportion. Mice infected with a 10 LD₅₀ dose of HSV-1 were treated with TJS-109 orally at doses of 1.25 to 20 mg/kg 2 days before, and 1 and 4 days after the infection. The treated groups had 80% (1.25 mg/kg), 40% (5 mg/kg) and 23% (20 mg/kg) mortality rates 25 days after the infection as compared with a 100% mortality rate in control mice treated with saline. When HSV-1 infected mice (recipients) received CD8⁺T cell fractions derived from spleens of mice treated with TJS-109 (donors), 70% of recipients survived, as compared with 0% survivors in the groups of mice treated with saline, B cell fractions, CD4⁺ T cell fractions or macrophage-enriched fractions prepared from the same donors. TJS-109 did not show any virucidal activities against HSV-1 or any virostatic activities on the growth of HSV-1 in Vero cells. These results suggest that TJS-109 protected mice exposed to lethal amounts of HSV-1 through the activation of CD8⁺ T cells.     

Suppression of the secondary immune response by specific antibody,when given together with the secondary antigenic stimulus

Summary It is generally believed that antibody-mediated immunosuppression can be only produced in non-primed individuals, and that this applies both to experimental animals and Rh-negative women at risk. However, in this paper it is reported that the additional injection of 0.2 ml of an antiserum to sheep erythrocytes (SE) together with a secondary antigenic stimulus of 10⁸ SE into mice, primarily immunized by a tiny dose of 5×10⁵ SE 28 days before, was capable of producing effective suppression of the secondary immune response.This work was supported by grants from the Deutsche Forschungsgemeinschaft.     

Ca-efflux,from direct membrane injury by CCl4, is elicited by amphiphilic vehicles in vitro

Direct membrane injury by CCl₄, in situations excluding metabolic activation, was evaluated in saponin-permeabilized hepatocytes and in microsomes by measuring immediate Ca²⁺ efflux. A good correlation appears between the Ca²⁺ efflux and the level of CCl₄ in the membrane and also the variations in fluidity. Mixtures of CCl₄ with water-soluble vehicles were used to improve the dispersion of CCl₄ in the medium. The mixtures varied in their ability to elicit the membrane effects of CCl₄. The performance of ethanol and, to a lesser degree, other alcohols, suggests the existence of a water stable structural organization between CCl₄ and these amphiphilic vehicles, facilitating the transfer of CCl₄ to the membrane.     

Bactericidal activity againstPseudomonas aeruginosa is acquired by cultured human monocyte-derived macrophages after uptake of myeloperoxidase

Myeloperoxidase (MPO) is an enzyme located within polymorphonuclear neutrophils capable of producting cytotoxic oxidant species that are particularly active against bacteria with polysaccharide capsules.Pseudomonas aeruginosa (10⁶ bacteria per 1 ml) are killed within 1 h in vitro by a MPO/H₂O₂/Cl^– system (48 mU=132 ng of MPO). The question arose as to whether human macrophages would acquire cytotoxic activity when loaded with this enzyme. Monocytes were therefore isolated from human blood and cultured for up to ten days to induce maturation to macrophages. These cells lost endogenous MPO within five days while H₂O₂ production in response to stimulation by phorbol myristate acetate (10^–6M) decreased to 23% within ten days. On the other hand, their capacity to take up exogenous MPO increased fourfold from day three to day ten. Human macrophages cultured from eight days (when both H₂O₂ production and MPO uptake were sufficient) were therefore used to study the effects of MPO uptake on cytocidal activity againstPseudomonas aeruginosa. After a 1 h MPO loading period, macrophages (5×10⁵ cells per ml) were incubated in the presence of bacteria (0.5 to 2×10⁶ bacteria per ml) for 2 h at 37°C. At a bacteria/macrophage ratio of 1, only 34.8±7.0% of bacteria survived (compared to killing by non-loaded macrophages), while 74.4±9.3% survived at a ratio of 4. From these results, we conclude that loading macrophages with exogenous MPO could enhance their microbicidal activity, suggesting a potentially useful therapeutic application.     

Collapsin response mediator protein-2 is a calmodulin-binding protein

Collapsin response mediator protein-2 (CRMP-2) plays a crucial role in axonal guidance and neurite outgrowth during neural development and regeneration. We have studied the interaction between calmodulin (CaM) and CRMP-2 and how Ca²⁺/CaM binding modulates the biological functions of CRMP-2. We have shown that CRMP-2 binds to CaM directly in a Ca²⁺-dependent manner. The CaM binding site of CRMP-2 is proposed to reside in the last helix of the folded domain, and in line with this, a synthesized peptide representing this helix bound to CaM. In addition, CaM binding inhibits a homotetrameric assembly of CRMP-2 and attenuates calpainmediated CRMP-2 proteolysis. Furthermore, a CaM antagonist reduces the number and length of process induced by CRMP-2 overexpression in HEK293 cells. Take together, our data suggest that CRMP-2 is a novel CaM-binding protein and that CaM binding may play an important role in regulating CRMP-2 functions. Received 26 June 2008; received after revision 18 November 2008; accepted 24 November 2008     

Metionina,etionina, colina nel metabolismo intermedio dei fibroblasti coltivatiin vitro

Summary The administration of ethionin to tissue cultures of fibroblastsin vitro causes a rapid and total inhibition of the growth of the transplanted tissues.It was observed that equimolar quantities of methionin are, on the contrary, well tolerated. The inhibition is therefore seen to be due to the dismetabolic activity of the antagonist.The contemporary administration of cholin is not able to restore the growth capacity of ethionin inhibited tissue cultures.In conclusion, ethionin produces a direct action on fibroblastsin vitro, probably intervening in the process of protein synthesis of the transplanted elements.     

Positive and negative inotropic effects of carbachol on the embryonic chick atrium

The effect of carbachol on twitch tension of atrial preparations from chick embryos of different incubation ages (3–14 days) was studied. At every age carbachol evoked negative (at low concentrations) and positive (at higher concentrations) inotropic responses. Maximal response values for both effects increased with age; in 3- and 5-day atria the positive inotropic response prevailed. The muscarinic antagonist pirenzepine inhibited the positive (on 5-day atria) and negative (on strips of 14-day atria) inotropic effects of carbachol with pA₂ values of 6.8 and 8.0, respectively, suggesting that muscarinic receptors mediating these effects belong to different receptor subtypes.