Search

Lectin-stimulated supernatants of human peripheral blood leukocytes (PBL) contained an activity that maintained the long-term growth of murine interleukin-3 (IL-3) dependent cell lines such as the 32 Dcl line. We compared this human IL-3 activity with the basophil promoting activity (BaPA) recently described by us. Besides lectin-stimulated PBL supernatants that contained both BaPa and IL-3, we found that BaPA was produced mainly by E rosette-positive T cells or the human Mo T cell line, while IL-3 was produced by human T hybridomas and, more interestingly, by human macrophage cell lines. Based on molecular weight (BaPA: 20-30 kilodaltons, IL-3: 15-19 kilodaltons) as well as on molecular charge (BaPA: pI = 6.1-7.5, IL-3: 4.9-6.2), we concluded that human IL-3 and BaPA are distinct factors.

In order to characterize the form of choline kinase in rat tissues, both electrophoretic and gel chromatographic patterns of choline kinase activity were compared in the liver, kidney, lung, whole intestine and carbon tetrachloride-induced liver cytosols. Kinetic parameters of the reaction were also compared for the main forms of choline kinase protein from these tissues. The overall results suggested strongly that choline kinase does not exist in one particular active form but exists in multiple forms in rat tissues. In the study present here, the electrophoretic patterns of both choline kinase and ethanolamine kinase activities were compared in rat liver, kidney, lung and intestinal cytosols. The results strongly supported the view that both kinase activities are represented on the same enzyme protein(s) in each of the rat tissues examined.

New evidence is provided that rat liver choline kinase exists in several distinct forms (choline kinases I, II and III) which differ in isoelectric point, molecular size and antigenicity against anti-rat kidney choline kinase IgG. Remarkable and selective induction of the choline kinase II and choline kinase III forms of choline kinase was caused similarly by the administration of polycyclic aromatic hydrocarbon carcinogen, 3-methylcholanthrene or hepatotoxic carbon tetrachloride (CCl4). The immunochemical approach further indicated that the elevation in the activity of choline kinase in the 3-methylcholanthrene- or CCl4-treated rat liver was not accompanied by a parallel increase in the amount of choline kinase II enzyme protein, compatible with the induction of either a small amount of new enzyme protein(s) with very high specific activity or another enzyme which might catalyze post-translational modification of choline kinase.

Transient myeloproliferative disorder accompanied by Down's syndrome has been characterized as exhibiting self-limiting haematological abnormalities. We studied six patients suffering from this disorder in order to clarify the biological nature of their blast cells. Metaphases of leucocytes stimulated with phytohaemagglutinin (PHA) showed trisomy 21 in all patients except one. The exception was constitutionally trisomy 21 mosaic (46,XY = 89/47,XY,+21 = 11). However, metaphases from the peripheral blood cells (blast cells: 70%) without PHA stimulation showed exclusively trisomy 21. Simultaneous examination for morphology and chromosomal analysis on single colonies revealed that granulocyte-macrophage (GM) colonies and an erythroid colony contained only cells with the trisomy 21 karyotype. The blast cells showed positive reactions for platelet peroxidase (PPO) and with monoclonal antibodies against platelet-megakaryocyte antigen (TP 80 and TP 82). In methylcellulose and liquid culture systems, high plating efficiencies were observed, and mainly mature basophils containing histamine developed in the presence of PHA-stimulated leucocyte conditioned medium (PHA-LCM). In vivo, mature neutrophils, basophils, eosinophils or megakaryocytes coexisted with PPO-positive blast cells in the peripheral blood of some patients with this disorder. These findings suggest that transient myeloproliferative disorder is characterized by a proliferation of PPO-positive blast cells with trisomy 21, although some heterogeneity may be seen.

We have established a new T-cell line (CL-8313) that produces interleukin-3 from the spleen cells of mice with a radiation-induced myeloproliferative disorder. IL-3 activity was detected at an extremely high level in the culture medium of the CL-8313 cell line in the absence of any exogenous stimulator. A large amount of IL-3 transcript also was demonstrated in CL-8313 cells. BPA- and CSF-activity was detected at a high level in the culture medium of the CL-8313 cell line. Southern blot analysis of high molecular weight DNA from the CL-8313 cells showed they had unique rearrangement of the antigen receptor beta chain gene. Therefore, we concluded that CL-8313 cells belonged to T-lineage lymphocytes and constitutively produced a high level of IL-3.

There are two phenotypically distinct subpopulations of mast cells in rodents: connective tissue-type mast cells (CTMC) and mucosal mast cells (MMC). These populations differ in their location, cell size, staining characteristics, ultrastructure, mediator content and T-cell dependency. Several investigators recently reported a further subclass of mast cells which arise when normal mouse haematopoietic cells are cultured with interleukin-3 (IL-3); IL-3 is an activity similar or identical to mast-cell growth factor, histamine-producing factor, or P-cell stimulating factor. These cultured mast cells are in many ways similar to MMC; they stain with Alcian blue but not safranin, contain chondroitin sulphate E proteoglycan rather than heparin proteoglycan and have relatively low histamine content, as do MMC. Although proliferation of MMC is known to be T-cell dependent in vivo and thought to be IL-3-dependent in vitro, the factors on which CTMC proliferation depends remain elusive. Here we show that mature CTMC purified from mouse peritoneal cells can proliferate in vitro in methylcellulose culture and maintain the appearance and function of CTMC. We also present evidence that mature CTMC cannot proliferate in the presence of pure IL-3 alone.

A case of an anaphylactic shock following topical application of chlorhexidine preparation is reported. Specific skin-sensitizing antibodies against chlorhexidine were demonstrated in the serum from the patient by a passive transfer test. IgE antibodies against chlorhexidine were also detected by radioallergosorbent technique (RAST). Paper discs conjugated with chlorhexidine-HSA (human serum albumin) significantly bound the IgE antibodies. Furthermore, all of the sera from seven other patients with shock reactions following the topical application of chlorhexidine preparation also showed high RAST counts. Both chlorhexidine gluconate and chlorguanide which represents approximately half a molecule of chlorhexidine inhibited the reaction in a dose-dependent fashion. It is suggested that the shock reactions following topical application of chlorhexidine are mediated by IgE antibodies against chlorhexidine and that chlorhexidine and chlorguanide share the same antigenic determinant.

Four commercially available penicillin preparations (benzylpenicillin, aminobenzylpenicillin, carbenicillin and sulbenicillin) and two kinds of the new semisynthetic penicillin preparations (mezlocillin and piperacillin) evoked systemic anaphylactic reactions in guinea pigs immunized with one of the following penicillin-Ascaris extract (As) conjugates in aluminum hydroxide gel (alum), i.e., benzylpenicilloyl (BPO)-As, aminobenzylpenicilloyl (ABPO)-As, carbenicilloyl (CBPO)-As, sulbenicilloyl (SBPO)-As, mezlocillin-As conjugate (MEZ-As) and piperacillin-As conjugate (PIP-As). The tracheal chains prepared from the tracheas of the guinea pigs immunized with BPO-As, MEZ-As, or PIP-As revealed anaphylactic contractions in vitro on addition of benzylpenicillin, mezlocillin, or piperacillin preparations. These results indicated that the semisynthetic penicillin preparations as well as benzylpenicillin preparation have enough antigenicity to evoke systemic anaphylactic reactions in guinea pigs immunized with penicillin-As conjugate. Such guinea pigs seem to be suitable animals for investigations of penicillin allergy.

All of the five commercially available benzylpenicillin preparations obtained from different sources and a PcG preparation prepared by filtration of a commercial PcG on Sephadex G10 elicited the systemic anaphylactic reactions in guinea pigs which had been immunized with benzylpenicilloyl (BPO)-Ascaris extract conjugate (BPO-As) mixed with aluminum bydroxide gel. These preparations could evoke no such reactions in guinea pigs immunized with BPO-bovine gamma globulin conjugate (BPO-BGG) emulsified with complete Freund's adjuvant. The severity of the systemic anaphylactic reactions correlated significantly with the titers of either 8-day passive cutaneous anaphylactic (8-day PCA) reactions or 4-hr PCA reactions evoked with the same benzylpenicillin preparations. In vitro benzylpenicillin preparation contracted the tracheas of the guinea pigs immunized with BPO-As. These results indicated that the commercially available benzylpenicillin preparations have enough antigenicity to evoke systemic anaphylactic reactions in guinea pigs immunized with BPO-As mixed with aluminum hydroxide gel. Such guinea pigs represent an animal model for investigation of penicillin allergy.

Our previous study indicated that the IgE antibody responses in mice immunized with intraperitoneal injection of the antigens mixed with diesel-exhaust particulates (DEP) were higher than those in the animals immunized with the antigens alone. We examined the adjuvant activity of DEP inoculated by the intranasal route, i.e., the natural entrance of DEP. In 3-week interval immunization, the IgE antibody responses in mice immunized with intranasal inoculation of ovalbumin (OA) mixed with DEP were higher than responses in the animals immunized with OA alone. DEP had an adjuvant activity for anti-OA IgE antibody production, even in a small dose such as 1 micrograms administered with a 3-week interval. Also in 1-week interval immunization, the enhancing effect of DEP on anti-OA IgE antibody production was demonstrated when mice were immunized with intranasal inoculation of OA and DEP. The possibility cannot be excluded that DEP, which are kept buoyant in the environmental atmosphere of urban districts, may exert an adjuvant activity for IgE antibody production after being inhaled into the human body and have some relation to the mechanism of the outbreak of allergic rhinitis caused by pollens in Japan.

A new kind of myeloproliferative disorder (L-8313) has been discovered. It was transplantable into syngeneic mice with spleen cells. The mice showed hepato-splenomegaly with a marked leukocytosis and anemia 3 weeks after transplantation of L-8313 cells. The number of GM-CFU and CFU-S per spleen increased to more than 40 times normal. The results of chromosomal and PGK analysis demonstrated that these increased stem cells were of host origin. Both the culture medium of the spleen cells and the serum from L-8313 bearing mice showed high levels of IL-3, BPA and CSF. Consequently, hematopoietic cells of the host mice underwent remarkable proliferation in response to these stimulating factors when L-8313 cells were transplanted. We also have been successful in establishing an in-vitro cell line and have maintained it for over one year. The phenotype of L-8313 cells was Thy 1.2 positive. Some L-8313 cells showed a positive acid phosphatase reaction but the cytochemical character of myeloid lineage was not observed. Therefore, L-8313 is considered to be a T-cell derived hematopoietic regulatory cell neoplasm with the ability to produce several hematopoietic stimulating factors.

We have been using 99mTc-RBC and 99mTc-HSA for cerebral angiography and measurement of the local cerebral blood volume (LCBV) and cerebral/peripheral hematocrit ratio (alpha). 99mTc-HSA, however, has a problem of poor stability in the blood. We conducted basic and clinical studies on 99mTc-HSA-D, a ready-to-use preparation with good stability in the blood. We measured the regional cerebral circulation dynamics in patients with cerebral diseases in comparison with 99mTc-RBC and 99mTc-HSA. 99mTc-HSA-D and 99mTc-RBC showed more than a 90% retention ratio in the blood 60 min after administration, and good SPECT images were obtained using these two pharmaceuticals. On the contrary, 99mTc-HSA was unstable and its retention ratio was 60s% 60 min after administration. 99mTc-HSA does not provide good SPECT images due to free 99mTcO4- which seems to be liberated from 99mTc-HSA in the body, etc. There were no adverse reactions or abnormal laboratory findings caused by the 99mTc-HSA-D administration in any of the patients. Based on these results, it is surmised that 99mTc-HSA-D is clinically useful for measurement of the cerebral blood volume and cerebral/peripheral hematocrit ratio (alpha).