Postjunctional α1- and α2-adrenoceptors mediating contraction in isolated human groin arteries and veins

By means of selective agonists and antagonists for α₁- and α₂-receptors, the α-receptor subtypes in human groin arteries and veins were characterized and compared. In the arteries the α₁-receptor blocker prazosin caused a concentration-dependent parallel displacement of the noradrenaline (NA) concentration-response (cr) curve without reduction of maximum (pA₂=9.86); the selective α₂-receptor antagonist rauwolscine in the concentration 10^-8 M caused a right-ward shift of the NA cr-curve without reduction of E_max, but 10^-7 M and 10^-6 M caused little or no further shift. In the veins, the two antagonists had the opposite effects. Rauwolscine caused a concentration-dependent right-ward shift of the NA cr-curve without depression of maximum (pA₂=9.03); prazosin 10^-9 M significantly displaced the NA cr-curve, whereas 10^-8 M and 10^-7 M caused little or no further shift. The responses to the α₂-receptor agonist clonidine in the arteries were too small to allow calculations of pEC50 values; in the veins contractions were elicited in all vessel segments investigated (pEC₅₀=6.24). Phenylephrine, selective for α₁-receptors, was significantly more potent in arteries than in veins. NA was significantly more potent in veins than in arteries. It is concluded that in human groin vessels, there is a functional predominance of arreceptors in the arteries and of a2-receptors in the veins.     

Influence of antimuscarinics on alpha-adrenoceptors in the female rabbit urethra

The effects of the tertiary amine atropine and its structural analogues homatropine and scopolamine, as well as the quarternary amine emeprone, were evaluated on noradrenaline (NA)-induced contractions of isolated female rabbit urethral ring preparations. In addition, the abilities of these antimuscarinics to inhibit ³H—dihydro—alpha—ergocryptine (³H—DHE) binding to alpha—adrenoceptors were studied on a crude membrane preparation from the female rabbit bladder base and urethra. Atropine and homatropine depressed the NA- induced contractions in a concentration-dependent way, whereas this was not seen with scopolamine. Emeprone 10^-5 10^-4 M augmented the contractions, an effect possibly attributable to a NA-uptake blocking effect. All antimuscarinics displaced specific ³H— DHE binding, the order of potency being atropine>homatropine>emeprone>scopolamine. In general a good correlation was seen between the binding and mechanical activity studies for atropine, homatropine and scopolamine, while this was not found for emeprone. It is concluded that alpha-adrenoceptor blockade by atropine can be observed at concentrations exceeding 10^-7 M. Scopolamine, showing alpha-adrenoceptor blocking properties only in high concentrations, may be used as an alternative for blockade of muscarinic cholinoceptors.     

Preponderance for either α or β-adrenoceptor mediated sensitization in the rat submaxillary gland

The sensitivity of the rat submaxillary gland was examined 3–4 weeks after either parasympathetic decentralization or sympathetic decentralization or denervation. The threshold doses for secretion of saliva of parasympathomimetic (methacholine) and sympathomimetic (noradrenaline, adrenaline, phenylephrine and isoprenaline) drugs were estimated and the amount of saliva secreted in response to supraliminal doses of these drugs was measured. Each type of operation caused the development of a supersensitivity that involved all three types of receptors, i.e. muscarinic cholinoceptors, alpha;-adrenoceptors and β-adrenoceptors. Following parasympathetic decentralization the sensitization was predominantly mediated via α-adrenoceptors, and also via cholinoceptors. Following sympathetic decentralization or denervation the postjunctional sensitization was predominantly mediated via β-adrenoceptors; most of the supersensitivity to noradrenaline, adrenaline and phenylephrine found after sympathetic denervation was of the prejunctional type. An increase in receptor density and an intracellular arrangement where the response of cholinoceptors and α-adrenoceptors is mediated via one pathway and the response of β-adrenoceptors via another are suggested as factors that may be of importance for the development of the postjunctional supersensitivity. The present study shows that the traffic of secretory impulses in the sympathetic nerve is of importance for the level of sensitivity of the secretory cells. Since postjunctional supersensitivity following sympathetic denervation did not exceed that following sympathetic decentralization it is suggested that under normal conditions a continuous release of noradrenaline from the nerve endings is of little importance for the level of sensitivity.     

Pharmacological characterization of postjunctional α-adrenoceptors in isolated human omental arteries and veins

The α-adrenoceptors in human omental arteries and veins were characterized and compared. In the arteries both prazosin (pA₂ 9.48) and rauwolscine (pA₂ 7.19) displaced the noradrenaline (NA) concentration-response (cr) curve towards higher concentrations without reduction of maximum. Neither clonidine, nor oxymetazoline had any consistent contractile effects. Phenylephrine had a lower potency than NA, but a similar intrinsic activity. In the veins, both prazosin (pA₂ 9.72) and rauwolscine (pA₂ 8.11) displaced the NA cr-curve towards higher concentrations, but also significantly depressed maximum. Clonidine and oxymetazoline contracted veins from 3 out of 7 and 4 out of 6 patients, respectively. Their pD₂-values were similar to that of NA, but their intrinsic activities were significantly lower. NA was more potent than phenylephrine in these vessels, and there was no significant difference in intrinsic activity. The results suggest that in human omental arteries, the postjunctional a-adrenoceptors are mainly of the α₁-type, even if a small population of α₂-adrenoceptors cannot be excluded. In omental veins, there seems to be a functionally important population of postjunctional α₂-adrenoceptors occurring together with a population of α₁-adrenoceptors.     

β1-and β2-adrenoceptor-mediated secretion of amylase from incubated rat parotid gland

The present in vitro investigation was undertaken in an attempt to obtain further information on β-adrenoceptor specificity and action in the rat parotid gland, with regard to amylase secretion. The β₁-selective agonist prenalterol was roughly 800 times more potent than the β₂-agonist terbutaline, and about 5 times more effective than noradrenaline in evoking amylase release. Propranolol was the most effective inhibitor of amylase release in all experiments. The β₁-selective antagonist metoprolol and H104/08 were also effective blockers of maximal noradrenaline-and prenalterol-induced release. The inhibition curves displayed biphasic shapes when amylase secretion was induced by noradrenaline, but not when prenalterol was the secretagogue. The β₂-antagonist H35/25 was without effect on maximal noradrenaline-and prenalterol-stimulated secretion. The amylase release evoked by submaximal concentration of terbutaline was inhibited by the two antagonists H35/25 and IPS 339. In another series of experiments propranolol and metoprolol clearly shifted the noradrenaline concentration-response curve to the right, whereas H35/25 was without effect. The results further demonstrate the major importance of the β₁-adrenoceptor (noradrenaline-activated) in eliciting amylase release from the rat parotid gland. However, it is also suggested that the β₂-adrenoceptors (terbutaline-activated) may to some extent serve the same function.     

β-adrenoceptor agonists do not cause tremor in the conscious rat

The ability of some β-adrenoceptor agonists to induce tremor via skeletal muscle β₂-adrenoceptors in conscious unrestrained rats has been investigated. Tremor was assessed by visual observation and by an objective method based on accelerometry. Infusion of isoprenaline or terbutaline did not cause tremor, neither did β-stimulation potentiate an established mild tremor produced by central muscarinic receptor stimulation. Since β-agonists readily produce tremor in man via skeletal muscle β₂-adrenoceptor stimulation, our findings indicate that these receptors have a different function in the rat.     

Molecular characterization and clinical presentation of HKαα and anti‐HKαα alleles in southern Chinese subjects

The HKαα allele is a rearrangement occurring in the α‐globin gene cluster containing both the ‐α^3.7 and ααα^anti4.2 unequal crossover junctions. The anti‐HKαα allele is the reciprocal product containing both the ‐α^4.2 and ααα^anti3.7 unequal crossover junctions, which had been predicted but had not been detected previously. The phenotypic feature and population frequency of these two unusual alleles were not described. We report the identification of nine individuals carrying the HKαα allele and two individuals carrying the anti‐HKαα allele in southern China and describe their phenotype and haplotype data. The molecular structures of HKαα allele and anti‐HKαα allele were confirmed by two‐round nested polymerase chain reaction assay. The mechanism of origin of both alleles is related to probably simultaneous double crossover. Heterozygotes of HKαα or anti‐HKαα allele show a normal hematological phenotype. Finally, we report the carrier rates of these both alleles in the Guangxi Zhuang Autonomous Region of southern China, namely, ∼0.07% for the HKαα allele and ∼0.02% for the anti‐HKαα allele.     

Acute blockade of β1-receptors in the asphyxiated sheep fetus

Acute blockade of β₁-receptors in the asphyxiated sheep fetus. Acta Physiol Scand 130 , 381–385. Received 5 November 1986, accepted 9 February 1987. ISSN 0001–6772. Department of Paediatrics, Landspitalinn, University Hospital, Reykjavik, Iceland and Department of Physiology and Department of Paediatrics I, University of Goteborg, Sweden. The effects of acute β₁-blockade on fetal cardiovascular reactions during asphyxia were evaluated in 11 exteriorized sheep fetuses. Gestational age was 110–142 days. Asphyxia was induced either by ventilating the mother with low oxygen gas mixture or by mechanical reduction of placental blood flow. During asphyxia all fetuses reacted to metoprolol injection with a decrease in heart rate, myocardial contractility, cardiac output and arterial blood pressure. Five experiments resulted in irreversible fetal cardiovascular collapse. Isoprenaline was given to the fetuses during hypoxia to test the ability to further increase heart rate and activate myocardial β-adrenoceptors. In those experiments with fetal cardiovascular demise after metoprolol, the isoprenaline injection did not result in a significant tachycardia. The surviving fetuses could increase their heart rate as a sign of a capacity to further increase the sympatho-adrenergic drive.     

αβ Lineage-committed thymocytes can be rescued by the γδ T cell receptor (TCR) in the absence of TCR β chain

Commitment of the αβ and γδ T cell lineages within the thymus has been studied in T cell receptor (TCR)-transgenic and TCR mutant murine strains. TCRγδ-transgenic or TCRβ knockout mice, both of which are unable to generate TCRαβ-positive T cells, develop phenotypically αβ-like thymocytes in significant proportions. We provide evidence that in the absence of functional TCRβ protein, the γδTCR can promote the development of αβ-like thymocytes, which, however, do not expand significantly and do not mature into γδ T cells. These results show that commitment to the αβ lineage can be determined independently of the isotype of the TCR, and suggest that αβ versus γδ T cell lineage commitment is principally regulated by mechanisms distinct from TCR-mediated selection. To accommodate our data and those reported previously on the effect of TCRγ and δ gene rearrangements on αβ T cell development, we propose a model in which lineage commitment occurs independently of TCR gene rearrangement.     

Liver distribution of γδ‐T‐cells in patients with chronic hepatitis of different etiology

The γδ T cells represent a minor unique T‐cell subpopulation long been considered as innate‐like immune cells. They are found in increased numbers in tissues from various inflammatory conditions. Their role in chronic hepatitis, however, is still discussed controversially. Fresh frozen tissues from 50 patients (18 cases hepatitis B infection, 25 hepatitis C, three cases with co‐infection of hepatitis B and C and four patients with autoimmune hepatitis) were investigated. Immunohistochemistry with primary antibodies detecting αβ and γδ TCR was used to evaluate their incidence and distribution in the different histological structures of the liver. The inflammatory infiltrate in all cases of chronic hepatitis was dominated by αβ T cells and was mainly localized in the portal tracts with formation of an interface hepatitis (95.3%αβ T cells; 4.7%γδ T cells). There were neither significant differences between inflammatory infiltrate nor the amount or percentage of γδ T cells between hepatitis B, C or autoimmune hepatitis. No accumulation of γδ T cells could be observed in cases of chronic hepatitis of different etiologies. The immune‐mediated phenomena in chronic hepatitis are dominated by αβ T cells. Thus, the adapted immune system is responsible for the inflammatory processes in chronic hepatitis.     

α-thalassaemia due to a single codon deletion in the α-1-globin gene. Computational structural analysis of the new α-chain variant

A new unstable α-globin chain associated with α-thalassemia phenotype has been found in a Spanish patient. Molecular analysis of the α-globin gene complex using PCR and non-radioactive single-strand conformation analysis, allowed to identify a new mutation in the second exon of the α-globin gene. Direct sequencing of the abnormal fragment revealed a 3 bp deletion, which led to the loss of a single codon corresponding to a Lys (K) residue at position 60 or 61 DK60 or DK61. Theoretical structural analysis, performed by computational methods, indicated that the loss of an amino acid residue at this position disturbed the contact region between the B and E-helices, affecting the overall stability of the molecule. Therefore, the DK60 or DK61 results in a structurally abnormal α-globin chain, not previously described, named Hb Clinic, which leads to the α-thalassemia phenotype in the heterozygote patient. No abnormal hemoglobin was detected by standard electrophoretic procedures, suggesting that this α-globin chain variant is so unstable that it may be catabolized immediately after its synthesis. This mutation was confirmed by PCR using an allele specific primer. Hum Mutat 11:412, 1998. © 1998 Wiley-Liss, Inc.     

Expression of HNF-1α and HNF-1β in various histological differentiations of hepatocellular carcinoma

Hepatic nuclear factor 1 (HNF-1) regulates genes in a hepatocyte-specific manner. It has been previously reported that the ratio of HNF-1α and HNF-1β mRNA is related to histological differentiation hepatocellular carcinoma (HCC). In this study, the expression levels of the HNF-1α and HNF-1β proteins were analysed relatively and quantitatively in various histologically differentiated HCC and surrounding non-cancerous tissues, and HNF-1α binding activity for the AT element of the B domain of the human α-fetoprotein enhancer was examined. Western blot analysis demonstrated that HNF-1α protein was expressed at a higher level in well-differentiated HCC tissues than in the surrounding non-HCC tissues; on the other hand, the HNF-1α protein was expressed at lower levels in moderately and poorly differentiated HCCs than in the surrounding non-HCC tissues. The levels of HNF-1β expression in well-differentiated and poorly differentiated HCCs were similar to and higher than those found in the respective surrounding non-cancerous portions. In binding assays, HNF-1 binding activity was high in well-differentiated HCC and lower in moderately and poorly differentiated HCCs. Most well-differentiated HCC cases showed immunohistochemical expression of HNF-1α. These findings show that poor histological differentiation of HCC correlates with decreases in the level and activity of HNF-1α proteins. © 1998 John Wiley & Sons, Ltd.     

Autoradiographic location of β-adrenoceptor subtypes in cat colon smooth muscle

In order to localize β-adrenoceptors ¹²⁵I-(—)pindolol (IPIN) was used in binding to sections from cat colon. The binding characteristics for IPIN to β-adrenoceptors on colon sections were estimated by demonstrating reversible binding in the presence of isoprenaline and by steroselective binding to the isomers of propranolol. The binding of IPIN to both β₁-and β₂-adrenoceptors was shown by biphasic displacement curves in the presence of the selective β-adrenoceptor compounds betaxolol, ICI 118.551 and procaterol. The colon sections were found to contain proportions of β₁-adrenoceptors (30–50%) and β₂-adrenoceptors (50–70%). In the autoradiographic studies, 100% of the developed grains after exposure of IPIN to the photographic emulsion were displaced by 50 μm of isoprenaline. By microscopic counting at autoradiographic grains, 30–40% of the grains were found in the circular smooth muscle, while 60–70% of the grains were found in the longitudinal smooth muscle. A concentration of 2 nm ICI 118.551 completely displaced all grains in the circular smooth muscle and partly displaced those found in the longitudinal smooth muscle. A high concentration of ICI 118.551 (1 μm ) displaced all grains above background from the smooth muscle. It is concluded that the circular smooth muscle only contains β₂-adrenoceptors, while longitudinal smooth muscle may contain a proportion of β₁-adrenoceptors. Whether such a location of β-adrenoceptors can be related to the β₁-adrenoceptor-mediated inhibition of colon motility can not be clarified from these studies. However, it seems that β₁-adrenoceptors are located to the longitudinal smooth muscle instead of to the myenteric plexus of the colon.     

γδ T cells are secondary participants in acute graft-versus-host reactions initiated by CD4+ αβT cells

To examine the role of T cell subpopulations in an acute graft-versus-host (GVH) reaction, γδ T cells and αβ T cells expressing one of the two prototypic Vβ gene families were negatively isolated from adult blood samples and injected into allogeneic chick embryos. CD4⁺ αβ T cells expressing either Vβ1 or Vβ2 receptors were equally capable of inducing acute GVH reactions, consistent with the idea that αβ T cell alloreactivity is determined by CDR3 variability. By themselves, the γδ T cells were incapable of inducing GVH reactions. However, host γδ T cells were recruited into the donor αβ T cell-initiated lesions, where they were activated and induced to proliferate. The data suggest that γβ T cells may play a secondary role in GVH reactions.     

VIP inhibition of vascular smooth muscle: complementary to β2-adrenoceptor mediated relaxation in the isolated rat portal vein

Exogenous VIP caused a concentration dependent inhibition of the spontaneous mechanical activity in the isolated rat mesenteric-portal vein preparation via a mechanism which was completely independent of the propranolol-blocked β-adrenoceptor, of high K⁺ in the medium and of exogenous bovine pancreatic polypeptide, neurotensin and opioids. The potency of VIP ((pD₂=7.52±0.18, n=6) was about 30 times higher than that of isoprenaline in the atropine and phentolamine-blocked preparation. The isoprenaline inhibition was mediated via a β₂-type of adrenoceptor with low apparent affinity for noradrenaline (intrinsic activity (a) = 0.27±0.01, n=8). Opposite effects of exogenous VIP and noradrenaline were on the other hand observed in the atropinized and β-blocked preparation. These results suggest that in the rat portal vein neuronal VIP and circulating adrenaline may be complementary in their antagonism of the α-adrenoceptor mediated increase in contractility.     

High frequency of CD8αα homodimer-bearing T cells in human fetal intestine

Immunohistochemistry on frozen sections was used to identify CD8αα cells and CD8αβ cells in human intestine. As observed previously, CD8αβ cells predominate (>95%) in tonsil and post-natal intestine. However in human fetal intestine (16–24 weeks gestation), almost half the CD8⁺ cells in the lamina propria are CD8αα, and many CD8αα cells can be identified in the epithelium. In contrast, in the T cell zones of the Peyer's patches, CD8αβ cells are dominant. The CD8αα cells are virtually all αβ T cell receptor positive. By analogy with the murine system, these CD8αα cells in the fetal gut may be directly derived from the marrow, undergoing thymus-independent differentiation in the gut mucosa.     

Expression of pro-inflammatory cytokines by TCRαβ+ T and TCRγδ+ T cells in an experimental model of colitis

An inflammatory bowel disease (IBD) comparable to human ulcerative colitis is induced upon transfer of T cell-depleted wild-type (F1) bone marrow into syngeneic T cell-deficient (tgε26) mice (F1 → tgε26). Previously we have shown that activated CD4⁺ T cells predominate in transplanted tgε26 mice, and adoptive transfer experiments verified the potential of these cells to cause disease in immunodeficient recipient mice. Using flow cytometry for the detection of intracellular cytokine expression, we demonstrate in the present study that large numbers of CD4⁺ and CD8⁺ TCRαβ⁺ T cells from the intraepithelial region and lamina propria of the colon of diseased, but not from disease-free mice, produced interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α). Large numbers of T cells from peripheral lymphoid tissues of these animals also expressed IFN-α and TNF-α, but few expressed interleukin-4, demonstrating g strong bias towards Th1-type T cell responses in these animals. TCRγδ⁺ T cells, typically minor constituents of the inflammatory infiltrate of the colon in F1 → tgε26 mice, also expressed IFN-γ at a high frequency upon CD3 stimulation. In light of these findings we examined the potential involvement of TCRγδ⁺ T cells by testing their ability to induce colitis in tgε26 mice. We report here that tgε26 mice transplanted with T cell-depleted bone marrow from TCRα^null and TCRβ^null animals developed IBD. Furthermore, disease in these mice correlated with the development of peripheral and colonic TCRαδ⁺ T cells capable of IFN-γ production. These results suggest that IFN-γ may be a common mediator of IBD utilized by pathogenic T cells of distinct phenotype.