T-cell epitope analysis of Mag 3, an important allergen from the house dust mite, Dermatophagoides farinae.

Here we describe the detection of T-cell epitope region on the house dust mite allergen Mag 3, which has been shown to trigger T-cell proliferation in mite-allergic asthmatic patients. We first examined murine T-cell epitope using T-cell fraction prepared from recombinant Mag 3 (r-Mag 3)-primed H-2k mice. Initial proliferation assay with truncated r-Mag 3 indicated that N-terminal 113 amino acid region was required for triggering T-cell activation.

Cloning and expression of Der f 6, a serine protease allergen from the house dust mite, Dermatophagoides farinae.

House dust mite allergen is thought to be a major cause of asthma. Characterization of these allergen molecules is therefore an important step for the development of effective diagnostic and therapeutic agents against mite-associated allergic disorders. Here we report molecular cloning and expression of the group 6 (chymotrypsin-like) allergen from the house dust mite, Dermatophagoides farinae.

[The proliferative response and immunoglobulin production in peripheral blood lymphocytes stimulated with Japanese cedar pollen antigens].

Peripheral blood mononuclear cells taken from 38 normal subjects and 50 Japanese cedar pollinosis patients were cultured with or without mitogens or Japanese cedar pollen antigens to study the proliferative response and immunoglobulin production in periphral blood lymphocytes. When stimulated with Con A or anti-CD 40 mAb, the proliferative responses in the patient group were significantly higher than those in the normal group, and when stimulated with anti-CD 40 mAb, the IgG 2 production was markedly dominant among the IgE and IgG subclass productions in both groups. When stimulated with Japanese cedar pollen antigens, the proliferative responses in the patient group were markedly augmented in comparison with the normal group, and showed a correlation with the serum anti-cedar IgE antibody titers in the patient group.

Altered antigenicity of M-177, a 177-kDa allergen from the house dust mite Dermatophagoides farinae, in stored extract.

Background: A high molecular weight allergen, M-177 (177 kDa) was isolated from Dermatophagoides farinae using a specific antibody raised to an allergenic clone Mag 3, which was obtained by immunoscreening a mite cDNA library. The potent IgE reactivity of M-177 is comparable with that of Der f 2.

Objective: The aim of this study was to analyse the molecular characteristics and the allergenic activity of M-177 in stored mite extracts.

Identification of T-cell epitope sequences on an important mite antigen.

Background: T-cell epitopes on Der 1 and Der 2 groups, the major mite allergens, have been intensively analysed, while those on the other important allergens remain to be elucidated. We have cloned four cDNAs coding for important mite allergens on the basis of frequency and capacity of IgE binding. Stimulatory action of glutathione S-transferase-fused Mag1 on lymphocytes from mite-allergic patients was relatively high among them.

Human T cell responses to recombinant mite antigens of Dermatophagoides farinae.

We studied T cell responses to four glutathione S transferase (GST)-fused mite antigens prepared in our laboratory using peripheral blood lymphocytes from mite-sensitive patients with bronchial asthma. Of the four recombinant antigens, purified GST-Mag3 had the strongest ability to cause patients' lymphocytes to proliferate, and its potency was almost comparable to that of crude mite bodies (Dfb) and faeces (Dff) antigens and a purified major antigen, Der f 2. The responder lymphocytes were mainly T cells, because the proliferative response was depleted by the treatment of lymphocytes with anti-CD3 antibody and complement, but not with anti-CD20 antibody and complement.

[Length of survival of elderly patients with chronic obstructive pulmonary disease].

We examined survival in 53 patients over 60 years old who had chronic obstructive pulmonary disease, and whose FEV1 was less than 60% of the forced vital capacity and was less than 60% of the predicted value. They comprised 34 men and 19 women. The mean age was 75.

[A comparison of asthma deaths and near-fatal asthma attacks].

This study compares the characteristics of fatal asthma patients and those of near-fatal asthma patients who had experienced severe life threatening attacks. Data from 67 fatal asthma patients and 80 near-fatal asthma patients were analyzed. The mean age of fatal asthma cases were older than near-fatal asthma cases, 51.

Cloning and characterization of a new allergen, Mag 3, from the house dust mite, Dermatophagoides farinae: cross-reactivity with high-molecular-weight allergen.

A new immunoreactive clone whose sequence is not homologous to that of any of the previously identified mite allergens was isolated by successive immunoscreening of a Dermatophagoides farinae cDNA library with rabbit antisera to an extract of the house dust mite and IgE in pooled sera from patients allergic to mites. Rabbit antibodies specific for the recombinant protein recognized a 177 kD protein in a mite body extract. This immunoreactive protein was located in the circumferential tissues of esophagus, gut and the other internal organs in mites.

B cell mitogenic activity of house dust mite, Dermatophagoides farinae, antigens.

The effect of mite antigens on murine and human lymphocytes was studied in vitro. Antigens prepared from Dermatophagoides farinae feces and bodies stimulated normal murine spleen cells to proliferate in a dose-dependent manner. The responder cells are B cells, because the response was reduced by the treatment of spleen cells with anti-immunoglobulin antibody and complement, but not with anti-Thy 1 antibody and complement.

Immunochemical characterization of recombinant and native tropomyosins as a new allergen from the house dust mite, Dermatophagoides farinae.

Background: Two predominant mite species, Dermatophagoides farinae and Dermatophagoides pteronyssinus, are responsible for the immediate hypersensitivity reaction. Although a large number of antigens and allergens was detected in a whole mite culture extract, only the major allergens of Der I and II groups have been extensively studied. Much information on other important allergens remains to be accumulated.

[Desensitization immunotherapy on patients with mite-positive bronchial asthma using purified mite feces antigen fractions].

At present, we are performing desensitization immunotherapy on patients with mite-positive bronchial asthma using purified mite feces antigen fractions. We obtained the following results in 13 patients 4 to 12 months after the start of treatment. 1) The fraction showing the maximum reaction in the skin test was HM1-2 (molecular weight: 150-155 kD) in two patients, HM2 (30-40 kD) in nine patients and HM3 (10-20 kD) in two patients, and immunotherapy was performed using these fractions.

High-molecular-size mite allergens with high carbohydrate content in mite extract containing mite feces of Dermatophagoides farinae.

A series of mite allergens with high carbohydrate content was fractionated from mite extract containing mite feces by ultrafiltration, and successive chromatography on Ultrogel AcA 54, Sepharose CL-6B, and DEAE Toyopearl 650S. The final fractions isolated were termed HM1-2-Eff, -0.1A, -0.

B cell mitogenic activity of sea squirt antigen.

The activity of sea squirt antigen, one of the allergy-inducing substances for humans, on murine and human lymphocytes was studied in vitro. Sea squirt antigen stimulated normal mouse spleen cells to proliferate, as detected by [3H]-TdR incorporation, in a dose-dependent manner. The responder cells are B cells because the response was reduced by the treatment of spleen cells with anti-immunoglobulin antibody and complement and passing through a nylon wool column, but not with anti-Thy-1 antibody and complement.

A low-molecular-size LM2 allergen in mite, Dermatophagoides farinae, extracts containing feces which elicits conjunctival reactions in patients with bronchial asthma.

A low-molecular-size antigen, LM2, which possesses a significant activity to elicit histamine release, was isolated from an ultrafilterable (Mr-cutoff < 10k) fraction of a mite extract containing feces by consecutive chromatography on Ultrogel AcA 54 and Sephadex G-25. The isolated antigen was still heterogeneous glycoproteins, giving a smeary band around pI 3-5 on an Ampholine PAG plate, with molecular weights of 6-8k and 4k on SDS-PAGE and Sephadex G-50 gel filtration, respectively. Chemical treatments of the antigen suggested that the epitope responsible for the elicitation of histamine release resided in the protein moiety.

Low-molecular-size allergens, LM-1s, in feces extract of Dermatophagoides farinae which elicit histamine release from washed blood cells of patients with bronchial asthma.

A significant activity to elicit histamine release was found in an ultrafilterable (Mr-cutoff < 10 kD) fraction of a mite feces extract from a spent mite medium. The activity was divided into two fractions (LM1 and LM2) on an Ultrogel AcA 54 column when monitored by histamine release assay using washed peripheral blood cells from mite-allergic patients. The larger-molecular-size antigen, LM1 was further separated into three allergenic fractions by consecutive chromatography on Sephadex G-50, GM-DEAE, and TSKgel ODS-120T.

Cloning and expression of cDNA coding for a new allergen from the house dust mite, Dermatophagoides farinae: homology with human heat shock cognate proteins in the heat shock protein 70 family.

An immunoreactive clone, which shares no homology with the major allergens, Der f I and Der f II, was isolated by screening of a Dermatophagoides farinae cDNA library with rabbit antiserum raised against an extract of the house dust mite and sera from patients with mite allergy. The deduced amino acid sequence of the cDNA of the clone is significantly homologous, up to 65.5%, to the carboxyl-terminal region of the heat shock cognate protein (hsc) 71 in the heat shock protein (hsp) 70 family.

Structure of IgE epitopes on a new 39-kD allergen molecule from the house dust mite, Dermatophagoides farinae.

Two IgE epitope sequences comprising Ser56-Pro-Val-Thr-Lys-Arg-Ala-Ser-Leu-Lys-Ile-Asp-Ser-Lys-Lys70 and Asp104-Val-Glu-Leu-Ser-Leu-Arg-Ser-Ser-Asp-Ile-Ala115 were identified by deletion analysis of the cDNA encoding a new 39-kD protein of mite allergen. A synthetic dodecapeptide corresponding to the latter epitope sequence functioned as a monovalent and mite-specific hapten. Replacement of each of the 12 amino acid residues with Gly, using site-directed mutagenesis, indicated that Arg110 may play a central role in IgE binding.

Cloning and characterization of cDNA coding for a new allergen from the house dust mite, Dermatophagoides farinae.

A cDNA clone encoding a new allergen from the Dermatophagoides farinae cDNA lambda gt11 library was isolated and sequenced. There was no amino acid sequence homology with other known allergens. The gene product, beta-galactosidase fusion protein, of the truncated cDNA on blot reacted with IgE in 13 of 43 sera from patients allergic to mites.

Differences in response of patient's blood cells with bronchial asthma to diverse house dust mite allergens by histamine release assay.

A HPLC method was developed for the determination of histamine released from washed blood cells of allergic patients after exposure to allergens in the presence or absence of plasma. The blood cells used were preserved in a vital state at 4 degrees C during at least 6 hours after blood-collecting. The responses of blood cells to an appropriately diluted series of a mite feces extract (Dff) solution were evaluated in three patient's groups (68 cases of hyposensitized, 59 cases of newly visited and 44 cases of the others) with bronchial asthma.

[Results of 29-year study of hoya (sea-squirt) asthma in Hatsukaichi, Hiroshima prefecture].

As remarkable improvements have been made to the working environment and working methods of oyster shucking workers during the past 30 years, a study was made on the effects of these improvements on hoya (sea-squirt) asthma and the following results were obtained. 1) The prevalence of hoya (sea-squirt) asthma among oyster shucking workers was 36.0%, 30.

[Immunotherapy using high concentration purified antigen showed remarkable effect in all cases].

Hoya (sea-squirt) asthma is a typical type I occupational asthma. A total of 22 females with this disease whose age ranged from 22 to 69 years were treated for two years with high concentration purified sea-squirt antigen named Ei-M having a molecular weight of 22,800. One case who developed ordinary bronchial asthma abandoned the immunotherapy during the early stage of the therapy, but in the remaining 21 cases remarkable effects were rapidly observed from the early stage of treatment.

Conjunctival allergenicity-supporting factor (CASF) in DIIIa as the sea squirt antigen capable of inducing asthmatic attack and conjunctival reaction in sea squirt asthma.

Among three glycoproteins, DIIIa, Ei-M, and Gi-rep, isolated from sea squirt as antigens capable of eliciting skin reaction specific to sea squirt allergy, only DIIIa induces asthmatic attack and conjunctival reaction, whereas Ei-M and Gi-rep do not. Periodate oxidation eliminated not only the conjunctival allergenicity but also the skin allergenicity from DIIIa, suggesting that the conjunctival and skin allergenicity could both be ascribed to the epitope residing in the carbohydrate chains of DIIIa. On the other hand, proteolysis with Pronase E and chemical modifications of the carboxyl or amino groups by various methods eliminated the conjunctival allergenicity from DIIIa, whereas the skin allergenicity was still retained.

Therapeutic effect and titers of the specific IgE and IgG antibodies in patients with sea squirt allergy (Hoya asthma) under a long-term hyposensitization with three sea squirt antigens.

Hyposensitization therapy with each of three sea squirt antigens, Gi-rep (molecular weight [MW] 106,000), Ei-M (MW 22,800), and DIIIa (MW 9980), have succeeded in 72%, 90%, and 36% of patients with sea squirt allergy, respectively, within the first year, and the effect has been maintained during subsequent 4-year maintenance therapy. All available sera from some of the hyposensitized patients were also examined for IgE and IgG titers against the most effective therapeutic antigen, Ei-M, and it was revealed that the Ei-M-specific IgG titer increased rapidly in the successfully hyposensitized patients, regardless of the therapeutic antigen used, except for a few patients. Furthermore, the high specific IgG titer, as well as the therapeutic effect, was maintained during the subsequent maintenance therapy.

[Laboratory and clinical studies of norfloxacin in respiratory tract infections].

Norfloxacin (NFLX), a new quinolone antibacterial agent, was investigated for its antibacterial activity and clinical efficacy on respiratory tract infections. The results obtained are summarized as follows: 1. Antibacterial activities were evaluated against 127 strains of various bacteria isolated from clinical sources.