Homocytotropic Antibody Research Articles

Experimental allergic conjunctivitis in inbred guinea pig strains with high respectively low bronchial allergic reactivity.

Inbred guinea pigs selected for high (IMM/S) respectively low (IMM/R) responsiveness to ovalbumin (OA) as measured by induced respiratory anaphylaxis, were investigated for atopic immune responses of their conjunctival mucosa. IMM/S animals sensitized either by inhalation of OA, or by instillation of antigen into the conjunctival sac, developed regularly an acute ocular inflammatory response to topical (conjunctival) challenge with the allergen. A minimum of 1 microgram OA dropped repeatedly into the conjunctival sac was enough for both ocular and systemic sensitization of the animals, but the minimal dose of effective challenge was considerably higher. In IMM/R animals, ocular hypersensitivity was not achieved by inhalation of OA, but after topical administration of the antigen some of the IMM/R strains could be challenged to ocular anaphylactic responses of the same intensity as observed in IMM/S animals. The conjunctivae of both IMM/S and IMM/R animals could be sensitized passively by intraperitoneal injection of guinea pig sera containing homocytotropic antibodies to OA, but topical administration of such sera had no effect.

Anaphylaxis-mediated epithelial Cl- secretion and parasite rejection in rat intestine.

Immunologically mediated expulsion of Trichinella spiralis infective larvae in the rat was used as a model to test the hypothesis that intestinal anaphylaxis induced by parasite antigen causes ion transport alterations in small intestinal epithelium, and that the small intestinal epithelium, by altering its physiologic state in response to mast cell-derived mediators, functions as an effector tissue in the expulsion process. Experimental results demonstrated that the rapid rejection response and antigen-inducible changes in net intestinal ion transport acquired through active immunization were transferable with serum containing a high titer of anti-trichinella homocytotropic antibody, as measured by the PCA test. Neither response was expressed in nonimmune hosts nor in recipients of serum in which the PCA-detectable antibody was reduced by heat treatment. Net ion transport by jejunal epithelium of both actively and passively immunized rats was measured in Ussing chambers by using the electrical correlate, short circuit current (Isc). Involvement of chloride secretion in antigen-induced alterations in Isc was deduced from the use of chemical agents that effectively and specifically blocked the antigen-induced Cl- secretory response. The results implicate anaphylaxis in both rapid worm rejection and altered epithelial ion transport.

Reaginic and homocytotropic IgG antibody response of Mastomys natalensis in experimental infections of filarial parasites (Litomosoides carinii, Dipetalonema viteae, Brugia malayi, B. pahangi).

Reaginic and homocytotropic IgG antibodies in sera using passive cutaneous anaphylaxis (PCA) test and antigen from Litomosoides carinii were followed in Mastomys natalensis, infected with L. carinii, Dipetalonema viteae, Brugia malayi or B. pahangi. Groups of animals with infections of various ages so as to cover a total infection period of up to 300 to 420 days post-infection (p.i.), depending on the species of parasites, were bled at 1- to 3-week intervals over periods of 50-112 days. In addition, intradermal tests were performed on animals infected with L. carinii to detect immediate type hypersensitivity. Reaginic antibodies were usually first detected in the 3rd week after infection. Thereafter, a marked increase of PCA titres was observed in the 4th week p.i. leading to maximum titres 4 weeks after infection with D. viteae and B. pahangi and 6 weeks after B. malayi infection. Mean maximum titres were between 1:40 and 1:160. Following the peak response, titres decreased markedly until the beginning of patency in infections with D. viteae, B. malayi and B. pahangi whereas a constant course was observed at this time in animals infected with L. carinii. A further rise in PCA titres occurred in all infections around the beginning of patency, resulting in maximum reagin levels in L. carinii infections (mean titre 1:80) and moderate titres in the other infections. During early patency there was an inverse relationship between microfilaraemia density and levels of reaginic antibodies. However, in the phase of decreasing parasitaemia in L. carinii infected animals, microfilariae counts and PCA titres were directly correlated. Homocytotropic IgG antibodies showed relatively constant PCA titres of about 1:20 in L. carinii infected Mastomys throughout the observation period. In D. viteae infections they were demonstrated at 30 days p.i., reaching titres of about 1:40. B. malayi infected animals showed a maximum titre of 1:40 40 days p.i.. Thereafter, titres decreased continuously and homocytotropic IgG antibodies were absent at 110 days p.i.. High titres were observed at day 150 but thereafter sera were negative. B. pahangi infected animals showed moderate titres (1:5) 35 days p.i.. Thereafter, antibodies were found at low titres until 115 days p.i.. Intradermal reactions in L. carinii infected animals generally increased in size from 30-60 but decreased when microfilariae appeared in the blood.(ABSTRACT TRUNCATED AT 400 WORDS)

Lung Nonresponsiveness Produced in Guinea Pigs by Aerosolized Antigen

Nonresponsiveness to aerosolized antigen induced by repeated exposure of guinea pigs to either bovine serum albumin (BSA) or ragweed (RW) aerosols was not antigen specific, since induction of lung sensitization with the same, as well as heterologous antigen, was prevented. However, prior lung sensitization to an antigen was maintained in spite of subsequent treatment with aerosolized antigen. In this case treatment with aerosolized antigen could abrogate sensitization of the lung to all subsequent antigens. Chopped lung pieces could not be passively sensitized by antisera to homologous or heterologous antigens unless they were previously treated at low pH. High levels of conventional and homocytotropic antibody were demonstrated in nonresponder animals. Nonresponsiveness induced by fed antigen, however, was antigen specific, and elevated homocytotropic antibody levels were not present. Aerosol-induced nonresponsiveness appears to be a local phenomenon which involves a blocking of the pulmonary mast cells by immune complexes or other blocking factors so that they can no longer respond to antigen challenge.

Potentiated IgE response in Nippostrongylus brasiliensis infested rats--sites of synthesis and traffic of cells secreting potentiated antibody.

The distribution and the kinetics of potentiated antibody synthesis have been studied at the cellular level in rats infested with Nippostrongylus brasiliensis using the homologous adoptive cutaneous anaphylaxis technique. In animals immunized in the hind footpads with alum-absorbed ovalbumin 10 days prior to infestation with the parasite, the major sites of potentiated anti-ovalbumin homocytotropic antibody synthesis were the regional lymph nodes of the gut and the lungs. Peyer's patches were weakly active late in the response and the spleen produced considerable amounts of potentiated antibody. The regional lymph nodes of the ovalbumin immunization sites were the only organs in which specific homocytotropic antibody synthesis was detected in uninfested control rats. The kinetics of synthesis of the potentiated antibody by cells correlated well with the levels of anti-ovalbumin IgE antibodies in the sera of the infested rats. A traffic of cells secreting anti-ovalbumin homocytotropic antibody was detected in the thoracic duct lymph, but not the mesenteric lymph of immunized uninfested rats. After infestation, the mesenteric lymph also contained cells secreting potentiated antibody. The mesenteric lymph is a major route by which IgE and potentiated IgE antibodies reach the circulation in infested rats. The possible mechanisms responsible for the effects of the parasite on antibody secretion in distant lymphoid organs are discussed.

Anterior hypothalamic lesions decrease anaphylactic contractions in guinea pig trachea in vitro by reducing histamine and LTC 4 reactivity

Discrete lesions in the anterior hypothalamus (AHA) of the guinea pig brain reduce the anaphylactic contraction of the trachea in vitro after active in vivo sensitization by 40%. This difference in anaphylactic contraction does not correlate with a difference in homocytotropic antibodies but coincides with a decreased smooth muscle response to the anaphylactic mediators histamine and leukotriene C 4. No difference in the β-adrenoceptor function of the tracheal preparations can be found. The results suggest that AHA lesions afford protection against anaphylaxis in actively sensitized guinea pigs at least in part through a reduced smooth muscle response to anaphylactic mediators.

Mouse IgE-mediated paw anaphylaxis in mice and rats.

Anaphylactic swelling is induced in the hind paws of mice and rats by intravenous challenge of antigen 72 h after subplantar injection of mouse IgE-rich antiserum into the paws, and can be quantified rapidly and accurately by measuring paw thickness. The edema model described may serve as an alternative to passive cutaneous anaphylaxis for studying immediate allergic reactions with mouse homocytotropic antibodies and screening potential antiallergic drugs.

Increases in airway reactivity to histamine and inflammatory cells in bronchoalveolar lavage after the late asthmatic response in an animal model.

An increase in airway reactivity after the late asthmatic response (LAR) has been noted in humans. Although these alterations in reactivity have been proposed to be associated with inflammation, no clinical study of the LAR has shown both increased airway reactivity and evidence of pulmonary inflammation. Employing an animal model of the LAR in rabbits developed in our laboratory (Am Rev Respir Dis 1982; 126:493-498), we examined changes in airway reactivity and pulmonary inflammation in rabbits having a late asthmatic response. Two groups of rabbits were studied: a control group (n = 10) received nonimmune serum, and a sensitized group (n = 10) received serum containing homocytotropic antibody (IgE) to ragweed extract (RWE) from rabbits previously immunized from birth with ragweed. Airway reactivity to histamine and the evaluation of different cell types in bronchoalveolar lavage fluid were determined 3 days before and 3 days after bronchial challenge with RWE in all rabbits. No control rabbit developed a LAR, and no significant changes occurred in this group's airway reactivity or cells in lavage fluid after bronchial challenge with RWE. In contrast, all sensitized rabbits developed a LAR, and airway reactivity for this group was markedly increased 3 days later. In addition, the total number of cells including both polymorphonuclear and mononuclear cells in lavage fluid increased significantly 3 days after the LAR. Ten days after the LAR in the sensitized group (n = 8), as airway reactivity returned towards baseline, so did the cells in lavage. We conclude that increased airway reactivity to histamine is temporally associated with pulmonary inflammation as defined by cells in lavage after the LAR in this model.

Modulatory Effects of Freund’s Adjuvant Treatment on Mast Cell Histamíne Release and Homocytotropic Antibody Synthesis

The present study examines the influence of Freund's complete adjuvant (FCA) injections on sensitized PVG rats with respect to serum levels of IgE and IgG2 alpha antibodies and total IgE (all assessed by radioimmunoassays) and the capacity of serosal mast cells to release histamine on challenge in vitro with 'immunological' secretagogues (specific antigen, anti-IgE, concanavalin A) or with compound 48/80. The rats were immunized with 10 micrograms ovalbumin (OA); alum, Bordetella pertussis vaccine, or silica gel were employed as adjuvants. Treatment with FCA was performed by single intraperitoneal injections 3, 2, or 1 week(s) before or 1 or 2 weeks after sensitization. Tests were conducted 3 weeks after sensitization. The results show that the effect of FCA treatment varied reproducibly with the adjuvant employed for sensitization and with the timing of the FCA administration. FCA treatment could either increase, fail to affect, or decrease total serum IgE and OA-IgG2 alpha antibody levels as well as serosal mast cell responsiveness, whereas OA-IgE antibody responses were decreased or not affected. Moreover, serum levels of OA-IgE and OA-IgG2 alpha antibodies and total IgE were affected by FCA treatment independently of each other. Finally, serosal mast cell responsiveness to a given secretagogue could be influenced by the FCA treatment apparently independently of that to other secretagogues. A salient finding was that effects of FCA treatment on mast cell responsiveness did not necessarily conform to effects on antibody synthesis. Collectively, these data support the opinion that the mechanisms of action of the IgE-promoting adjuvants employed differ and suggest that the expression of serosal mast cell responsiveness to each examined secretagogue can be regulated separately. They also suggest that the serosal mast cell sensitizing capacity of homocytotropic antibodies may not be adequately quantified by immunochemical methods employing reagents prepared against IgE and IgG2 alpha protein.

Vernal conjunctivitis. Model studies in guinea pigs immunized topically with fluoresceinyl ovalbumin.

Repeated topical applications of fluoresceinyl ovalbumin (FL-OA) to the conjunctival sac of guinea pigs sensitized them for conjunctival type 1 hypersensitivity reactions and mast cell degranulation. Guinea pigs infected with Ascaris suum, with high titers of circulating anti-A suum IgE and IgG1 antibody, also produced conjunctival type 1 reactions on topical challenge with A suum antigen. These reactions were no more intense than those of animals topically sensitized and challenged with FL-OA, which in some instances had no detectable serum homocytotropic antibody. Persistently reactive animals that had undergone repeated type 1 conjunctival reactions had histological findings (eg, papillary changes with extensive epithelial eosinophil infiltrates, epithelial thickening or thinning, numerous goblet cells, subepithelial lymphoid cell infiltrates, and new vessel formation) resembling those of human atopic vernal conjunctivitis.

Mouse paw anaphylaxis

Anaphylactic swelling passively transferred to the hind paws of mice with homologous antibodies was investigated. The ascites fluid containing homocytotropic antibodies of the IgG1 class in a high concentration was obtained by repeated injections of bovine serum albumin in complete Freund's adjuvant into the peritoneal cavity of female ICR mice. The standard procedure for mouse paw anaphylaxis defined was as follows: female ICR mice received subcutaneously 25 μl of a 1:20 diluted ascites in the hind footpad and 2 h later were given intravenously 1.0 mg of the antigen in saline, and subsequently at 5 min intervals paw thickness was assessed by means of a simple thickness gauge. Maximal paw edema occurred invariably 15 min after antigen challenge. Sex differene in response was not observed. Heating at 56°C for 4 h resulted in a reduction of the activity of mouse ascites by approximately 40%. Histological examinations revealed that paw anaphylaxis was accompanied by mast cell degranulation and release of biologically active constituents from mast cell granules. Promethazine, pyrilamine, diphenhydramine, methysergide, cyproheptadine, ketotifen and phenoxybenzamine provided significant protection from the anaphylactic reaction in mice, while bromocriptine, dexamethasone and indomethacin were inactive. Relative drug effectiveness in paw anaphylaxis, passive cutaneous anaphylaxis and active and passive systemic anaphylaxis in mice was compared and discussed. Consequently, it was shown that mouse paw anaphylaxis was not only convenient for quantitative assessment of homocytotropic antibodies, but also useful in he routine work for screeningof potential anti-allergic drugs.

Changes in bronchial anaphylactic reactivity induced in guinea-pigs by long-term treatment with histamine H2-agents.

Guinea-pigs were sensitized to ovalbumin (OA) by immunization regimens chosen to cause antigen-induced bronchial anaphylactic responses mediated mainly either by IgE-like antibodies or by IgG1-like antibodies. Treatment of the IgE-producing animals for three weeks with the histamine H2-receptor antagonist cimetidine (1 mg kg-1 i.p. once a day) or with the H2-agonist dimaprit (0.1, 1.0, or 10 mg kg-1 i.p. once a day) led to a significantly reduced bronchial response capacity compared with that of the saline-treated controls challenged intravenously with antigen one week after the end of treatment. The changes were biphasic and not strictly dose-dependent. In contrast, acute treatment of immunized animals with a single dose of cimetidine (10 or 30 mg kg-1 i.v.) or dimaprit (1 or 10 mg kg-1 i.v.) 2 min before challenge with OA did not significantly affect the bronchial anaphylactic response. However, long-term treatment with cimetidine (10 mg kg-1) or the dimaprit analogue, S-[4-(N, N-dimethylamino)-butyl] isothiourea (SKF Compound 91488) (1 mg kg-1), which is reported not to activate H2-receptors, had no effect on the response capacity. Treatment with cimetidine (1 mg kg-1) or dimaprit (1 mg kg-1) did not influence the response capacity to antigen challenge in IgG1- type animals. Dimaprit (1 mg kg-1) did not affect the responsiveness to intravenous provocation with histamine in 'IgE-type' animals. Antigen-induced release of histamine from chopped lung tissue in vitro was not significantly affected in 'IgE-type' animals treated with cimetidine (1 mg kg-1) or dimaprit (1 mg kg-1). Treatment of immunized animals with cimetidine or dimaprit one week before and one week after a booster injection of antigen also led to reduced response capacity compared with that of saline-treated controls. However, the serum levels of IgE-like homocytotropic antibodies of these animals were not reduced; on the contrary, those of IgG1-antibody were increased in dimaprit-treated animals. These data show that intermittent treatment with histamine H2-agents reduces reagin-mediated anaphylactic response capacity in vivo in actively sensitized guinea-pigs by an as yet undefined mode of action.

Role of IgE in fascioliasis

A prominent phenomenon of helminth infections is their stimulation of a substantial IgE response, both serum total IgE and parasite specific IgE, and there is now considerable evidence that the latter is involved in protective immunity to helminths: Parasite specific IgE has been shown as an essential factor in antibody dependent cell-mediated cytotoxicity reactions (ADCC) of various trematodes and nematodes in vitro. In Fasciola hepatica infections however, there is only limited information about the mechanisms of ADCC and the possible role of specific IgE in the immunity to this parasite. Rat eosinophils have been shown to adhere and to degranulate onto newly excysted F. hepatica in vitro in the presence of immune serum, while no bound complement was detectable on the worms. Also the killing capacity of eosinophil major basic protein was demonstrated but the particular involvement of reaginic antibodies has not yet been cleared. Time course studies in experimentally F. hepatica infected calves showed that the highest levels of homocytotropic antibodies were reached between 20 and 28 weeks of infection which coincides with the period of the parasite expulsion from the bile ducts of the host. This finding, together with the massive accumulation of mast cells at the bile ducts and the liberation of histamine from blood-cells of infected cattle when contacted with fluke antigen, further suggests that IgE might play a role in the defence mechanism of the host against F. hepatica infections. Using radioimmunoassay techniques in the rat model the increase of F. hepatica ES-specifc IgE is coincident with the increase of total serum IgE and the response shows a biphasic pattern. The first peak reflects the migratory phase of young flukes through the liver parenchyma, whereas the second increase appears when adult flukes establish in the bile ducts. However, the significance of these reactions is not yet understood and further investigations might possibly show wether circulating IgE can perform some regulatory mechanisms in immunity to liver fluke infections. In addition, it has been found that F. hepatica infections in rats — similarly to other helminth infections — were able to potentiate an unrelated IgE response. They also induced a prolonged elevation of serum total IgE, which declined after anthelmintic treatment. Other experiments suggested that the serum total IgE in F. hepatica infections is parasite-dose dependent. However, these observations are still not cleared and need to be further investigated.

A reagin-like antibody in horse serum. II. Anti-human IgE induced reversed cutaneous anaphylaxis-like responses in horse skin.

Fc specific anti-human IgE serum induced prolonged reversed cutaneous anaphylaxis (RCA)-like reactions in horse skin. Morphologically and histologically, these reactions resembled passively induced late cutaneous anaphylaxis responses in human skin, but differed from reversed passive Arthus responses induced in horse skin using anti-horse IgG serum. The induction of RCA-like responses in horse skin by anti-human IgE indicates shared Fc antigenic determinants on human IgE and a horse homocytotropic or reagin-like antibody.

The effect of cyclophosphamide on autoimmune response to rats immunized with modified accessory glands

It was found that the organ specific hemagglutinating autoantibodies to rat male accessory glands can be suppressed by the injection of a single dose of cyclophosphamide applied 3 days after the first immunization. On the other hand, injection of the drug 3 days before immunization did not modify the response, that is, there were comparable incidence and titers in both treated and the control animals. Cyclophosphamide did not appear to act on the homocytotropic antibodies. These results indicate that cyclophosphamide acts only when it is administered after antigenic stimulation, to suppress hemagglutinating antibodies production.

112 Immunization of guinea pigs to produce heatlabile homocytotropic antibody

112 Immunization of guinea pigs to produce heatlabile homocytotropic antibody

Dissociation between cutaneous basophil hypersensitivity and basophil sensitization.

We investigated the antigen-specific sensitization of basophils in the peripheral circulation of guinea pigs using an in vitro histamine release assay. Animals were primed for cutaneous basophil hypersensitivity (CBH) by intradermal injection of either keyhole limpet hemocyanin (KLH) or ovalbumin (OA) in saline. Basophils isolated from these animals 7 days after sensitization did not release histamine to the specific antigen in vitro. On day 9, however, basophils did release histamine when challenged with the antigen. These results were in contrast to 24-hr skin tests that showed a typical CBH profile both on day 7 and day 9. When animals were immunized via the footpad by emulsification of antigen in incomplete Freund's adjuvant, basophil sensitization (i.e., histamine release) either occurred earlier (day 7, KLH) or at lower antigen concentrations (OA). Thus, dissociation of in vivo CBH and in vitro basophil sensitization was demonstrated. The lack of correlation was only seen early after immunization and depended on the route of immunization used. An additional finding was a transient blood basophilia that also was dependent on the route of immunization, the antigen used, and the time post-sensitization. Our results are consistent with the hypothesis that at early times post-immunization, CBH is mediated by T cells and not via homocytotropic antibodies, but that at later times such antibodies may also play a role in the elicitation of CBH reactions.

Protective effects of the glucocorticoid, budesonide, on lung anaphylaxis in actively sensitized guinea-pigs: inhibition of IgE-but not of IgG-mediated anaphylaxis.

1 The effect of glucocorticoid pretreatment on antigen-induced bronchoconstriction was studied in guinea-pigs actively sensitized to two different ovalbumin regiments (one producing IgE- and IgG-like antibodies and the other exclusively IgG-like antibodies). 2 Budesonide (50 mg/kg) and hydrocortisone (50 mg/kg) given as one intraperitoneal injection 15-20 h before and anaphylactic tests or as two consecutive intraperitoneal injections 5 and 6 days before, led to a decreased bronchial capacity. In this respect glucocorticoid pretreatment was effective only in guinea-pigs sensitized to produce both IgE-like and IgG-like antibodies. 3 Budesonide pretreatment also reduced the capacity of anaphylactically-challenged chopped lung tissue to release histamine in guinea-pigs sensitized to produce both IgE- and IgG-like antibodies. 4 Budesonide pretreatment did not change the levels of circulating IgG1a and IgE-like homocytotropic antibodies as measured by passive cutaneous anaphylaxis; nor did it affect histamine or methacholine-induced bronchoconstriction in vivo or the capacity of histamine or methacholine to contract the guinea-pig isolated trachea preparation of the isoprenaline-induced relaxation of this preparation. 5 The selective inhibitory effects of budesonide and hydrocortisone on IgE-mediated but not IgG-mediated anaphylaxis and the relevance to human atopic disease are discussed.

Antigen-induced bronchial anaphylaxis in actively sensitized SD rats. Effects of immunization and provocation doses of antigen and of pretreatment with DSCG, theophylline, terbutaline and a new anti-allergic xanthine derivative, D4026.

Bronchial anaphylactic reactions, estimated as increase in intratracheal pressure, were precipitated by intravenous injections of antigen into actively sensitized SD rats. The degree of bronchial reactivity was found to depend on both the challenge dose and the immunization dose of antigen; therefore the course of the capacity to respond was recorded as a function of these variables. The degree of the bronchial anaphylactic response could be reduced by pretreatment with disodium cromoglycate, terbutaline or a new anti-allergic xanthine derivative, D 4026, in some groups of animals. The efficacy of each agent was found to depend on the dose of antigen used for sensitization and for provocation of the bronchial reaction rather than on the strength of the response. Taken together, the data suggest that more than one type of homocytotropic antibody mediates bronchial anaphylactic reactivity in the SD rat.

Animal model for testing human Ascaris allergens

Guinea pigs immunized through the nasal route by an antigenic preparation of humanAscaris lumbricoides produced predominantly homocytotropic antibodies. The sensitization of the homologous skin required a latency period. Hypersensitivity reaction was triggered within 30 min. of the antigen challenge. The antibodies were sensitive to heat and β-mercaptoethanol treatments and appeared to be similar to the IgE type of immunoglobulins of man and rabbit. The antigenic preparation elicited an immediate type of skin hypersensitivity reaction also in human subjects harbouring the Ascaris parasite. The guinea pig model appears suitable for testing the activity of Ascaris allergens.