Hajime Tsunoo

Acute cadmium intoxication in inbred mice: A study on strain differences

Male C3H, BALB/c and DBA/2 mice were injected subcutaneously with a single dose of 30 mumol CdCl2/kg. Most of the C3H mice died within 15 h, while all of DBA/2 survived for at least 48 h. At 6 h after Cd-injection, histology revealed severe hemorrhage, fatty change, zonal hepatocyte necrosis and congestion of C3H livers, while these findings were rare in the other strains even at 48 h post-injection. The rate of metallothionein (MT) induction at 6 h was significantly lower in C3H mice than in the other two strains.

Identification of human T cell epitopes in Japanese cypress pollen allergen, Cha o 1, elucidates the intrinsic mechanism of cross‐allergenicity between Cha o 1 and Cry j 1, the major allergen of Japanese cedar pollen, at the T cell level

Background Pollens from species of Cupressaceae family are one of the most important causes of respiratory allergies worldwide. In Japan, many patients with pollinosis have specific IgE to both pollens of Japanese cypress (Chamaecyparis obtusa) and Japanese cedar (Cryptomeria japonica). The sequences between Cha o 1 and Cry j 1, the major allergens of Japanese cypress and Japanese cedar pollens, respectively, are 80% identical.

Genetic influence on induction of metallothionein and mortality from cadmium intoxication

Abstract Male mice of C3H, BALB/c, and DBA/2 strains were injected subcutaneously with a single dose of 30 μmol of CdCl2/kg. Within 48 h after administration, the mortality of C3H, BALB/c, and DBA/2 was 7 9 , 2 7 , and 0 8 , respectively. At 6 h after administration, severe damage was observed in livers of C3H, but no remarkable change was observed in livers of BALB/c or DBA/2. In C3H mice, rate of hepatic metallothionein synthesis was significantly lower than in BALB/c or DBA/2. From these results, it is clarified that the mortality from Cd-intoxication is determined by the amounts of hepatic metallothionein the induction of which depends on genetic predisposition.

Human placental ferritin receptor

Brush-border membranes from human placenta were prepared and their purity was clarified by biochemical and morphological methods. Ferritin binding to these prepared membranes was examined using horse spleen 125I-apoferritin, and was found to be completed within 10 min at 37 degrees C and pH 7.5. The amount of ferritin bound to the membranes was found to be proportional to the amount of membrane added and saturable for a given amount of the membrane in the presence of excess ligand. The membranes exhibited specific ferritin binding with a Ka of 2.3 X 10(7) M-1 at pH 7.5. A competitive binding assay indicated that horse spleen 125I-apoferritin binding was inhibited by a 10-fold molar excess of horse spleen ferric ferritin and a 500-fold molar excess of human transferrin. These results suggest that human placental brush-border membranes have specific receptors for horse spleen apoferritin molecules.

Primary Structure of Mouse Liver Metallothionein -I and -II

Primary structure of metallothionein from several animal species has been determined or is under investigation in several laboratories. This paper reports isolation, chemical properties and amino acid sequence of mouse liver metallothionein -I and -II.

Peptide Specificity, HLA Class II Restriction, and T–Cell Subsets of the T–Cell Clones Specific to Either Cry j 1 or Cry j 2, the Major Allergens of Japanese Cedar (Cryptomeria japonica) Pollen

Background: Cry j 1 and Cry j 2 are thought to be the major allergens of Japanese cedar pollen. HLA class II types capable of presenting T–cell epitopes in both allergens and their role in induction of T–cell subsets are not well known. Methods: CD4+ T (Th)–cell clones (TCCs) specific to either Cry j 1 or Cry j 2 were generated. HLA class II restrictions were determined by their reactivity to the T–cell epitope in the presence of antigen presenting cells sharing matched types. Interleukin (IL)–2, interferon–γ, IL–4, and IL–5 contents in the supernatants of TCCs were estimated using enzyme immunoassay. Results: Peripheral blood mononuclear cells (PBMC) from patients induced proliferation with 100 μg/ml Cry j 1 or 3–10 μg/ml rCry j 2 stimulation. T–cell epitopes in Cry j 1 were presented to Th cells by the gene products of DRA1*01/DRB1*0901, DRA1*01/DRB5*0101, DQA1* 0102/DQB1*0602, and DPA1*01/DPB1*0501; those in Cry j 2 were restricted by DRA1*01/DRB1*0901, DRA1* 01/DRB1*1501, DRA1*01/DRB4*01, DRA1*01/DRB5* 0101, DQA1*0102/DQB1*0602, DPA1*01/DPB1*0201, and DPA1*01 and *0202/DPB1*0501. Type 2–like cells were preferentially induced in Cry j 1 stimulation, while an almost equal number of type 2– and type 1–like cells was induced in rCry j 2. Conclusions: No clear correlation existed between peptide specificity, HLA class II restriction and induction of Th–cell subsets, suggesting that the requirement of different dose of Cry j 1 or Cry j 2 to induce proliferation in PBMC may lead to distinguishable difference in induction of Th subsets between TCCs specific to Cry j 1 and Cry j 2.

An immunomodulatory protein, ling ZHI-8, facilitates cellular interaction through modulation of adhesion molecules

Ling Zhi-8 (LZ-8), a novel immunomodulatory protein, markedly enhanced the expression of CD11b, but not CD11a, CD13, CD14, CD18, CD33 or HLA-DR, on the U937 cell line in a dose-dependent fashion. It also induced ICAM-1 expression on vascular endothelial cells and significantly augmented gamma - interferon-induced cellular binding between vascular endothelial cells and U937. Furthermore, LZ-8 increased the expression of CD2, but not VLA4, VLA5 or LFA3, on MOLT4 and enhanced rosette formation between human T cells and sheep red blood cells. These data suggest that LZ-8 exerts its pharmacological effect by modulating adhesion molecules on immunocompetent cells.

Recognition of T Cell Epitopes Unique to Cha o 2, the Major Allergen in Japanese Cypress Pollen, in Allergic Patients Cross-Reactive to Japanese Cedar and Japanese Cypress Pollen

BACKGROUND Pollens from species of the Cupressaceae family are one of the most important causes of respiratory allergies worldwide. Many patients with pollinosis have specific IgE to both allergens from Japanese cedar and Japanese cypress pollen. We set out to identify T cell epitopes in Cha o 2, the second major allergen of Japanese cypress pollen. METHODS T cell lines (TCL) and T cell clones (TCC) specific to Cha o 2 were generated from allergic patients cross-reactive to Japanese cedar and Japanese cypress pollen. T cell epitopes in Cha o 2 were identified by responses of TCL stimulated with overlapping peptides. Abilities of IL-4/IFN-gamma production by TCC were evaluated using enzyme immunoassay. RESULTS Using TCL, 11 dominant and subdominant T cell epitopes were identified in Cha o 2. The subsets of TCC were predominantly of T helper 2-type. A T cell epitope p141-160 in Cha o 2 and corresponding peptide in Cry j 2 showed high homology. Although TCC PC.205.159 responded to stimulation with p141-160 in Cha o 2, it did not respond with corresponding peptide in Cry j 2, therefore, the T cell epitope was unique to Cha o 2. CONCLUSIONS Eleven T cell epitopes that were identified are unique to Cha o 2. Cha o 2 is a putative aeroallergen that can potentially sensitize human T cells. We concluded that generation of T cells specific to Cha o 2 in allergic patients acts as one of the causes of continuous allergic symptoms in April.

A receptor for formaldehyde-treated serum albumin on human placental brush-border membrane

Formaldehyde-treated serum albumin (f-Alb) is known to be taken up and degraded by sinusoidal liver cells via receptor-mediated endocytosis. We report that 125I-labeled f-Alb (125I-f-Alb) binding to human placental brush-border membranes also occurs. This binding reached equilibrium within 40 min at 37 degrees C. Kinetic studies demonstrated the presence of saturable binding with an apparent Kd of 2.1 micrograms of f-Alb/ml and a maximal binding of 2.3 micrograms/mg of membrane protein at pH 7.5. Maximal binding was observed at between pH 7.5 and 8.0. 125I-f-Alb binding to the membranes was little inhibited by a 1000-fold molar excess of ovalbumin, human apo-transferrin and native bovine serum albumin. No binding was observed with membranes which had been pretreated with proteinase or trypsin. This f-Alb receptor was extremely heat-stable, since the binding was not abolished even by pretreatment of the membranes at 78 degrees C for 30 min. EDTA, Ca2+ and Mg/4 had no effect on 125I-f-Alb binding, so the binding was independent of divalent cations. These data suggest that a receptor specific for f-Alb exists on human placental brush-border membranes of syncytial trophoblasts.

A ligand-receptor binding assay by receptor immobilization

Using transferrin-transferrin receptor binding as a model of ligand-receptor binding, we have developed a new and simple binding assay for the solubilized receptor. Solubilized membrane proteins containing transferrin receptor were immobilized by covalent binding to beads having chemical reactive epoxide groups, and then 125I-labeled transferrin was added to the beads. Dose-dependent, ligand-specific, and saturable binding of 125I-labeled transferrin to the immobilized membrane proteins were demonstrated and a Scatchard analysis derived affinity of Kd = 1.8 X 10(-9) M was obtained. These results indicate that the immobilization of receptors onto beads may be useful in a simple binding assay of the solubilized receptor.