1.
Greiff, Lennart, et al.
(författare)
Effects of topical platelet activating factor on the guinea-pig tracheobronchial mucosa in vivo
1997
Ingår i: Acta Physiologica Scandinavica. - 0001-6772. ; 160:4, s. 387-391
Tidskriftsartikel (refereegranskat) abstract
Platelet activating factor (PAF) has been reported to produce a variety of airway effects including epithelial damage and increased airway-lung absorption of hydrophilic tracers. The present study examines effects of PAF on the guinea-pig tracheobronchial mucosa in vivo. Vehicle with and without PAF (4.0 and 8.0 nmol) was superfused onto the tracheobronchial mucosa. The levels of 125I-albumin, previously given intravenously, were determined in tracheobronchial lavage fluids as an index of mucosal exudation of plasma. The mucosa was also examined by scanning electron microscopy. In separate animals, 99mTc-DTPA (a low molecular weight, 492 Da, hydrophilic tracer) was superfused onto the mucosal surface through an oro-tracheal catheter, together with vehicle or PAF (8.0 nmol). A gamma camera determined the disappearance rate of 99mTc-DTPA from the airways as an index of mucosal absorption. PAF produced dose-dependent mucosal exudation of plasma up to 20-fold greater than control (P < 0.001). However, PAF did not damage the epithelium and the absorption ability of the airway mucosa was unaffected. The results, in contrast to previous reports, suggest that PAF may not readily damage the airway mucosa even at large exudative doses of the agent. The present finding support the view that the plasticity of the epithelial junctions allows the creation of valve-like paracellular pathways for unidirectional clearance of extravasated plasma into the airway lumen. We suggest that endogenous PAF may participate in first line respiratory defence reactions by causing lumenal entry of bulk plasma without harming the epithelium.
2.
Korsgren, Magnus, et al.
(författare)
Allergic eosinophil-rich inflammation develops in lungs and airways of B cell-deficient mice
1997
Ingår i: Journal of Experimental Medicine. - 1540-9538. ; 185:5, s. 885-892
Tidskriftsartikel (refereegranskat) abstract
Immunoglobulins (Ig), particularly IgE, are believed to be crucially involved in the pathogenesis of asthma and, equally, in allergic models of the disease. To validate this paradigm we examined homozygous mutant C57BL/6 mice, which are B cell deficient, lacking all Ig. Mice were immunized intraperitoneally with 10 micrograms ovalbumin (OVA) plus alum, followed by daily (day 14-20) 30 min exposures to OVA aerosol (OVA/OVA group). Three control groups were run: OVA intraperitoneally plus saline (SAL) aerosol (OVA/SAL group); saline intraperitoneally plus saline aerosol; saline intraperitoneally plus OVA aerosol (n = 6-7). Lung and large airway tissues obtained 24 h after the last OVA or SAL exposure were examined by light microscopy and transmission electron microscopy (TEM). The Ig-deficient mice receiving OVA/ OVA treatment had swollen and discolored lungs and exhibited marked eosinophilia both in large airway subepithelial tissue (49.2 +/- 12.0 cells/mm basement membrane [BM] versus OVA/ SAL control 1.2 +/- 0.3 cells/mm BM; P < 0.001), and perivascularly and peribronchially in the lung (49.3 +/- 9.0 cells/unit area versus OVA/SAL control 2.6 +/- 0.6 cells/unit area; P < 0.001). The eosinophilia extended to the regional lymph nodes. TEM confirmed the subepithelial and perivascular localization of eosinophils. Mucus cells in large airway epithelium increased from 1.5 +/- 0.8 (OVA/SAL mice) to 39.5 +/- 5.7 cells/mm BM in OVA/OVA treated mice (P < 0.001). OVA/SAL mice never differed from the other control groups. Corresponding experiments in wild-type mice (n = 6-7 in each group) showed qualitatively similar but less pronounced eosinophil and mucus cell changes. Macrophages and CD4+ T cells increased in lungs of all OVA/OVA-treated mice. Mast cell number did not differ but degranulation was detected only in OVA/OVA-treated wild-type mice. Immunization to OVA followed by OVA challenges thus cause eosinophil-rich inflammation in airways and lungs of mice without involvement of B cells and Ig.
3.
Persson, Carl, et al.
(författare)
The mouse trap
1997
Ingår i: Trends in Pharmacological Sciences. - 0165-6147. ; 18:12, s. 465-467
Tidskriftsartikel (refereegranskat) abstract
Mouse models of asthma are now being used extensively in drug research. However, the successful unravelling of combinatorial interplays of cells and molecules in the murine airways may not be matched by equally successful demonstrations of an asthma-like pathophysiology. Here, Carl Persson, Jonas Erjefalt, Magnus Korsgren and Frank Sundler discuss the fact that major features of asthma may still need to be demonstrated in the airways of allergic mice.
4.
Elies, Rozenn, et al.
(författare)
Immunochemical and functional characterization of an agonist-like monoclonal antibody against the M2 acetylcholine receptor.
1998
Ingår i: European journal of biochemistry / FEBS. - 0014-2956. ; 251:3, s. 659-66
Tidskriftsartikel (refereegranskat) abstract
Monoclonal antibodies were raised against a peptide corresponding to the second extracellular loop of the M2 acetylcholine receptor. One of the monoclonal antibodies, B8E5, was selected for further characterization on the basis of its high yield, its isotype (IgG2a), its dissociation kinetics and its agonist-like activity. The epitope recognized by B8E5 corresponded to the N-terminal part of the second extracellular loop of the receptor (V-R-T-V-E-) as determined by competition immunoassays and epitope scanning. The KA of B8E5 for the target peptide was assessed by surface plasmon resonance (SPR) to be 6.5x10(7) M(-1) by equilibrium and 3.7x10(7) M(-1) by kinetic analysis. B8E5 recognized the M2 acetylcholine receptor on rat cardiac tissue. It only recognized the non-reduced receptor in immunoblots. The antibody had no effect on antagonist binding but decreased the affinity for the agonist carbachol. B8E5 decreased the beating frequency of neonatal rat cardiomyocytes. The effect was specific since it was blocked by the target peptide and the antagonist atropine. The EC50 of the antibody corresponded to the KA measured by surface plasmon resonance. The physiological effect of the antibody did not lead to desensitization. The Fab fragments had no physiological effect; subsequent addition of anti-mouse IgG however restored the physiological effect. These results confirm that the N-terminus of the second extracellular loop is a functional target for antibodies against the M2 acetylcholine receptor. They suggest that the functional epitope is only accessible in the non-reduced receptor. The antibodies act through a functional dimerization of the receptor.
5.
Fu, Michael, 1963, et al.
(författare)
Agonist-like activity of antibodies to angiotensin II receptor subtype 1 (AT1) from rats immunized with AT1 receptor peptide.
1999
Ingår i: Blood pressure. - 0803-7051. ; 8:5-6, s. 317-24
Tidskriftsartikel (refereegranskat) abstract
In the present study, rats were immunized with angiotensin II receptor subtype 1 (AT1) receptor peptides for 3 months to see if the immunization produced specific anti-AT1 receptor antibodies and if continuous stimulation for 3 months affected blood pressure or induced morphological changes in the organs containing AT1 receptors. Our results showed that there were constant high levels of circulating antibodies throughout the study period in all rats of the immunized group, but not in the control rats, and that there were almost no significant cross-reactions of antisera with AT2 receptor peptide and alpha1 adrenoceptor peptide, except in four rats, which showed low cross-reactions with alpha1 adrenoceptor and AT2 receptor peptides. When an affinity-purified anti-AT1 receptor antibody was used, it specifically displayed the AT1-stimulatory positive chronotropic effect and also localized AT1 receptors. However, in the immunized group, saturation binding of AT1 in homogenates from kidneys showed no difference either in maximal binding sites (Bmax) or in antagonist affinity (Kd). No difference in mRNA of AT1a was found in either kidney or heart, and no morphological changes in the organs were observed, as compared with the control group. Furthermore, immunization did not cause hypertension. In conclusion, the synthetic peptide corresponding to the second extra-cellular loop of the human AT1 receptor was able to produce highly specific and functionally active anti-AT1 receptor antibodies, but unable to induce pathological structural changes or hypertension.
6.
Fu, Michael, 1963, et al.
(författare)
Immunohistochemical localization of angiotensin II receptors (AT1) in the heart with anti-peptide antibodies showing a positive chronotropic effect.
1998
Ingår i: Receptors & channels. - 1060-6823. ; 6:2, s. 99-111
Tidskriftsartikel (refereegranskat) abstract
Antibodies were produced against a synthetic peptide corresponding to amino acids (165-191) of the second extracellular loop of the human angiotensin II receptor subtype 1 (AT1) in rabbits. The purified antibodies had an apparent affinity of about 1 nM and were monospecific for the AT1-receptor peptide. Chemical modification of the carboxyl groups (glu at positions 173 and 185) and the sulfhydryl group (cys at position 180) of the AT1-receptor peptide did not alter the relative affinity of the coated AT1-receptor peptide to antibodies. The antibodies specifically stained CHO cells expressing the rat AT1a receptor. Immunoblots on rat kidney revealed that the antibody recognized a protein band of 59 +/- 3 kDa in a dose-dependent manner and this band was no longer detected after preincubating the antibodies with AT1-receptor peptide. Using electron microscopic and immunofluorescence immunocytochemistry techniques, angiotensin II receptors were detected in (1) the sarcolemma, T-tubules and nuclei of rat cardiomyocytes, (2) the transluminal side of endothelial cells and (3) fibroblast cells. These localizations are specific, as the immunostaining did not appear when preimmune rabbit serum was used and was blocked after preincubating antibodies with antigenic peptide. Functionally, these antibodies did not affect the ligand binding properties of the receptors but displayed agonist-like activity as shown by dose-dependent increases in beating frequency in cultured neonatal cardiomyocytes. These results suggest that the antibodies against the second extracellular loop of human AT1 receptors were able to specifically recognize AT1 receptors. In addition, they extend the observation that the second extracellular loop of the G-protein coupled membrane receptors is a specific target for antibodies with agonist-like activity.
7.
Holtbäck, U, et al.
(författare)
Receptor recruitment: a mechanism for interactions between G protein-coupled receptors.
1999
Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424. ; 96:13, s. 7271-5
Tidskriftsartikel (refereegranskat) abstract
There is a great deal of evidence for synergistic interactions between G protein-coupled signal transduction pathways in various tissues. As two specific examples, the potent effects of the biogenic amines norepinephrine and dopamine on sodium transporters and natriuresis can be modulated by neuropeptide Y and atrial natriuretic peptide, respectively. Here, we report, using a renal epithelial cell line, that both types of modulation involve recruitment of receptors from the interior of the cell to the plasma membrane. The results indicate that recruitment of G protein-coupled receptors may be a ubiquitous mechanism for receptor sensitization and may play a role in the modulation of signal transduction comparable to that of the well established phenomenon of receptor endocytosis and desensitization.
8.
Liu, H R, et al.
(författare)
Screening of serum autoantibodies to cardiac beta1-adrenoceptors and M2-muscarinic acetylcholine receptors in 408 healthy subjects of varying ages.
1999
Ingår i: Autoimmunity. - 0891-6934. ; 29:1, s. 43-51
Tidskriftsartikel (refereegranskat) abstract
Autoantibodies to cardiac beta1-adrenoceptors and M2-muscarinic receptors have mainly been found in the sera of patients with idiopathic dilated cardiomyopathy (DCM). In order to elucidate the pathological significance of these autoantibodies in DCM, it is necessary to understand their characteristic distribution in a healthy population of different genders and ages. The peptides corresponding to the sequences of the second extracellular loops of the human beta1-adrenoceptor and M2-muscarinic receptors were therefore used as antigens to screen the sera of 408 healthy subjects of different ages (ranging from 0.5 to 85 years). Of 408 sera, 41 (10.0%) and 46 (11.3%) recognized the beta1-adrenoceptor and M2-muscarinic receptor peptides respectively. Of the positive sera for beta1-adrenoceptors and M2-muscarinic receptors, up to 63.4% and 56.5% had both anti-beta1-adrenoceptor and anti-M2-muscarinic receptor autoantibodies respectively. The antibody titres of the positive sera of healthy subjects were all of a low level, with a geometric mean titre of 1:42+/-1.9 for anti-beta1-adrenoceptor antibodies and 1:51+/-1.7 for anti-M2-muscarinic receptor antibodies. The frequency of occurrence of autoantibodies to both receptors in the sera of healthy subjects increased significantly with age. In conclusion, the autoantibodies to beta1-adrenoceptors and M2-muscarinic receptors in the sera of healthy subjects are characterized by a low frequency of occurrence and low titre, with the frequency of occurrence increasing with age.
9.
Matsui, S, et al.
(författare)
Active immunization of combined beta1-adrenoceptor and M2-muscarinic receptor peptides induces cardiac hypertrophy in rabbits.
1999
Ingår i: Journal of cardiac failure. - 1071-9164. ; 5:3, s. 246-54
Tidskriftsartikel (refereegranskat) abstract
BACKGROUND: The high prevalence of patients with dilated cardiomyopathy (DCM) with anti-beta1-adrenoceptor and/or anti-M2-muscarinic receptor autoantibodies in their sera has been observed. However, the pathophysiological role of these autoantibodies in the development of cardiomyopathy is unknown. We previously reported an experimental model of early-stage DCM-like cardiomyopathy induced by immunizing rabbits for 1 year with synthetic peptides corresponding to the sequence of the second extracellular loop of either beta1-adrenoceptor or M2-muscarinic receptor. Because approximately half the sera of patients with DCM that recognize one of the two receptor sequences also recognize the second sequence, a model was created in rabbits simultaneously immunized with the synthetic peptides corresponding to the second extracellular loop of the beta1-adrenoceptor and M2-muscarinic receptor. METHODS AND RESULTS: All rabbits (n = 8) immunized with both peptides had a high titer of both anti-beta1-adrenoceptor and anti-M2-muscarinic receptor autoantibodies in their sera, whereas none of the sera from control rabbits injected with saline (n = 9) was positive. No significant cross-reaction with peptides other than those used for immunization was found. The weight of the hearts of immunized rabbits increased significantly. The hearts of immunized rabbits showed marked concentric left ventricular hypertrophy with mild inflammatory cell infiltration. In these rabbits, mild or moderate interstitial fibrosis was also observed. In electron micrographs, immunized rabbits showed focal myofibrillar lysis, loss of myofilament, and a marked increase in the number of mitochondria and deposition of dense granules in both sarcoplasm and myofibrils. Conversely, one of the control rabbits showed scant mononuclear cell infiltration. However, in this control rabbit, no significant alteration was found by electron microscopy. CONCLUSION: Our results showed the coexistence of both anti-beta1-adrenoceptor and anti-M2-muscarinic receptor autoantibodies in the sera has pathophysiological importance, shown by their ability to induce cardiac hypertrophy in rabbits.
10.
Matsui, S, et al.
(författare)
Myocardial injury due to G-protein coupled receptor-autoimmunity.
1998
Ingår i: Japanese heart journal. - 0021-4868. ; 39:3, s. 261-74
Tidskriftsartikel (refereegranskat) abstract
One of the main mechanisms for dilated cardiomyopathy is likely to be autoimmune mediated myocardial damage. So far, a variety of autoantibodies have been detected against a number of putative autoantigens in the sera of patients with dilated cardiomyopathy. A growing body of studies have confirmed that autoantibodies against the second extracellular loop of beta 1-adrenoceptors and M2-muscarinic receptor are present in 30-40% of patients with dilated cardiomyopathy. These anti-beta 1-adrenoceptor and anti-M2-muscarinic receptor antibodies can not only decrease the binding sites of antagonist but also recognize the target receptors. Moreover, these two autoantibodies possess an 'agonist-like' stimulatory effect on the target receptors. In order to elucidate whether the autoantibodies against these autoimmune epitopes play an important role in the pathogenesis of dilated cardiomyopathy, we immunized rabbits over a period of one year with synthetic peptides corresponding to the second extracellular loop of the beta 1-adrenoceptor and the M2-muscarinic receptor. These peptides induced morphological changes in the heart similar to those found in dilated cardiomyopathy. These clinical and experimental findings suggest that these receptor autoantigens are of pathogenic importance in the development of dilated cardiomyopathy in vivo.
