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Heating of 7S globulin alone did not cause precipitation, but addition of lIS globulin to the system did. Gel filtration and ion exchange chromatography, followed by electrophoresis of the precipitate and supernatant, suggested an interaction through disulfide bonding between the 7S and 11 S globulin subunits. )"'he basic and f3 subunits tended to be located in the precipitate by interaction through a secondary force on heating. On the other hand, the acidic and (X(X' subunits tended to be located in the supernatant by interaction through disulfide bonding on heating.Using the predictive parameter of amino acids, each macroscopic parameter of the soybean globulin subunits was calculated on the basis of their amino acid composition and a relationship with their behavior on heating is discussed. 1229Heating contributes to the orientation of texture in many food systems.! --4) For example, .precipitation and heat-setting in hot water form the filament in a spinning process,5) and when making tofu, a traditional Japanese food, the heating of soybean milk plays a major role in forming its characteristic gel, while heating tofu to high temperature produces "aburage" (fried tofu) as textured proteins. 6 ) To acquire a basic knowledge about the thermal aggregation behaviour of food proteins, soybean lIS globulin has been used as a model for the examination of thermal aggregation, 7 --9) because it contains the sulfbydryl and disulfide residues, which playa major role in the formation of texture. In a food system, not only lIS globulin, but also 7S globulin must contribute to the orientation of texture by interacting with each other on heating. Thus, as a model system for heating food proteins, a mixture of purified 7S and lIS globulins was settled upon for this study.In this present investigation, we have studied heat-aggregated and -dissociated products by means of gel filtration, ion exchange chromatography and electrophoresis. MATERIALS AND METHODSMaterials. Soybean, cv. Raiden, was used throughout this work. Defatted meal was prepared from the soybean by grinding, defatting with n-hexane and removing the solvent at room temperature. All reagents were of the highest grade. Sepharose CL-6B and CL-2B, and DEAESephadex A-50 were obtained from Pharmacia Fine Chemicals.Preparation of 78 and 118 globulins. Thanh's crude 7S globulin 1 0) was prepared and purified by the method described in the previous paper. 11) The concentration of 7S globulin in the standard buffer was determined spectrophotometrically, using the value E~r~= 5.74. 12 ) Purified 11 S globulin was prepared by the method described in the previous paper. 7 ) Each protein was adjusted to a 1%concentration.Procedure for heat treatment. An equal mixture solution of 1% 7S and lIS globulins was used for heat treatment. Heating studies and experiments with 0.02 M Nethylmaleimide (NEM) were carried out by the method described in the previous paper. 9 ) The heated solutions were centrifuged for 10 min at 5000 rpm if a precipitate was present.Electrophoresis. Acetic ...

Efferocytosis, which is the homeostatic phagocytosis of apoptotic cells, prevents the release of toxic intracellular contents and subsequent tissue damage. Impairment of efferocytosis was reported in alveolar macrophages (AMs) of patients with chronic obstructive pulmonary disease (COPD), a common disease caused by smoking. In COPD, histone deacetylase (HDAC) activity is reduced in AMs. We investigated whether the reduction of HDAC activity is associated with the impairment of efferocytosis. Murine AMs were collected by bronchoalveolar lavage and their ability to efferocytose apoptotic human polymorphonuclear leukocytes was assessed. Pre-treatment of AMs with cigarette smoke extract (CSE) or trichostatin A (TSA), an HDAC inhibitor, suppressed efferocytosis and CSE reduced HDAC activity. TSA inhibited the activity of Rac, a key mediator of efferocytosis. These TSA-induced impairments were restored by treatment of AMs with aminophylline, a potent activator of HDAC. To further elucidate the underlying mechanism, we explored a role of CD9 in TSA-induced impairment of efferocytosis. CD9 is a transmembrane protein of the tetraspanin family that facilitates the uptake of several pathogens and other material. TSA profoundly down-regulated the expression of CD9 on AMs. The expression of CD9 was partly down-regulated by the Rac inhibitor. Pretreatment with an anti-CD9 mAb or CD9 small interfering RNA inhibited efferocytosis, which was attributable to the reduced binding of AMs to apoptotic cells. These results suggest that smoking impairs efferocytosis via inhibition of HDAC/Rac/CD9 pathways. Aminophylline/theophylline is effective in restoring the impairment of efferocytosis and might have benefit for the treatment of patients with COPD.

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