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The transport of Fe(2+) and other divalent transition metal ions across the intestinal brush border membrane (BBM) was investigated using brush border membrane vesicles (BBMVs) as a model. This transport is an energy-independent, protein-mediated process. The divalent metal ion transporter of the BBM is a spanning protein, very likely a protein channel, that senses the phase transition of the BBM, as indicated by a break in the Arrhenius plot. The transporter has a broad substrate range that includes Mn(2+), Fe(2+), Co(2+), Ni(2+), Cu(2+), and Zn(2+). Under physiological conditions the transport of divalent metal ions is proton-coupled, leading to the acidification of the internal cavity of BBMVs. The divalent metal ion transporter can be solubilized in excess detergent (30 mM diheptanoylphosphatidylcholine or 1% Triton X-100) and reconstituted into an artificial membrane system by detergent removal. The reconstituted membrane system showed metal ion transport characteristics similar to those of the original BBMVs. The properties of the protein described here closely resemble those of the proton-coupled divalent cation transporter (DCT1, Nramp2) described by, Nature. 388:482-488). We may conclude that a protein of the Nramp family is present in the BBM, facilitating the transport of Fe(2+) and other divalent transition metal ions.

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Rabbit articular cartilage does not secrete cathepsin B in organ culture. By established methods for modulating the chondrocyte phenotype in vitro, however, the synthesis, intracellular storage, and secretion of cathepsin B were followed up over a period of two months. With chondrocytes grown in monolayer cultures both the intracellular pool of the enzyme and its secretion were very small initially, but increased progressively to a factor of 110 after eight weeks. The secretion of cathepsin B was strongly depressed after transferring the cells from monolayer to collagen gel cultures. In contrast, collagenase was secreted in almost the same amounts during the whole period in both monolayer and collagen gel cultures. The cells cultured in collagen gels secreted more collagenase than those grown in monolayers. The reversible switch of cathepsin B secretion suggests that this enzyme, unlike collagenase, is a marker of the dedifferentiated chondrocyte phenotype. Cathepsin B was localised within cultured chondrocytes using antibodies raised against rabbit liver cathepsin B and shared with it many catalytic properties. Its Mr, however, was higher (34,000 compared with 27,000) and showed an unusual resistance to denaturation at neutral-alkaline pH, which may confer on this enzyme an important role in the degradation of cartilage matrix.

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Organ cultures of explanted V2 carcinoma specimens as well as cultured V2 carcinoma cells produced a cytokine which stimulated rabbit skin fibroblasts to synthesize increased amounts of cathepsin B. The cytokine was released by the tumor cells as a heterogeneous family of polypeptides: two inactive forms (Mr = 55,000 and 68,000) which could be activated by limited proteolysis with trypsin and three active forms with Mr values of 12,000, 16,000 and 18,000. The treatment of inactive cytokine-containing tumor-conditioned media with trypsin, followed by chromatographic separation of the products, suggested that the high-Mr inactive components may represent precursors of the active forms. Cathepsin B was immunolocalized in the tumor-host interzone in co-cultures of tumor and host tissues. Some other possible activities of the tumor cytokine which emerged from previous studies, such as the induction of host cells to produce increased levels of collagenase and extracellular matrix, as well as the stimulation of host cell proliferation, are discussed in the light of the new findings and are proposed as an important mechanism in tumor invasion.

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Rabbit V2 carcinoma cells and normal rabbit skin fibroblasts produced cysteine proteinases with properties similar to those of purified rabbit liver cathepsin B. Both cell types secreted into the culture medium enzymes with an apparent Mr of 43,000, which reacted with synthetic substrates commonly used for cathepsin B. After limited proteolysis with pepsin or treatment at pH 3, the Mr = 43,000 species could be converted into forms with Mr = 34,000 and an increased specific activity. In the intracellular pool of both V2 carcinoma cells and fibroblasts, a cysteine proteinase with the same Mr of cathepsin B (27,000) was found. Despite the similarity in molecular size, substrate specificity and sensitivity to inhibitors, the tumor and fibroblast enzymes were not identical in their stability at pH greater than or equal to 7 and were produced by the 2 cell types in considerably different amounts. In terms of enzyme units and normalized to an equal cell number, the ratios of fibroblast enzyme/tumor enzyme were as follows: secreted 130-150; intracellular, 150-180. The pH stability of the cysteine proteinases was determined quantitatively by measuring the half-life of enzyme activity. At pH 8.0 and 25 degrees C the secreted tumor cysteine proteinase had a half-life of at least 5 hr, whereas the secreted fibroblast enzyme and liver cathepsin B had half-lives of 8.8 min and 4.4 min, respectively.

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Cathepsin G, the chymotrypsin-like serine proteinase from human polymorphonuclear leucocytes, cleaves human IgG. The relative susceptibilities of the four IgG subclasses to the action of this enzyme were studied kinetically and showed the following decreasing order of susceptibility: IgG3 much greater than IgG4 greater than IgG1 greater than IgG2. IgG1 and IgG2 produced primarily F(ab')2 and traces of Fc-related fragments. IgG4 gave rise to both Fab and F(ab')2 as major products, and small amounts of an Fc-related fragment were detected. The cleavage of IgG3 produced various fragments, depending on the experimental conditions: The primary fragments were Fab and Fch (Fc covalently joined to the extended hinge-region polypeptide of IgG3) and an intermediate Fab-Fch species. Both Fab and Fch were further degraded by cathepsin G. Fch was gradually split, giving rise to three subfragments that were finally degraded to dialysable peptides. The enzyme further cleaved the Fab fragment in the heavy-chain portion and released a polypeptide probably representing the VH domain.

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Cathepsin G, the chymotrypsin-like enzyme from human polymorphonuclear leukocytes, cleaves human IgM and produces two major fragments that closely resemble those released by leukocyte elastase digestion of IgM. An F(ab)2 mu-like fragment, mol. wt 140,000, retains some reactivity with an anti-Fc mu antiserum and is antigenically deficient with respect to the IgM subunit. The other fragment is an Fab mu-like product with a mol. wt of 54,000. Both cathepsin G fragments are indistinguishable from the elastase counterparts by immunochemical analysis. An Fc mu fragment could not be recovered. The kinetic course of the cleavage shows that cathepsin G produces Fab mu fragments at a higher rate than F(ab)2 mu, whereas the contrary is valid for elastase. Beside the two major fragments and low mol. wt peptides, cathepsin G releases also a product with the same mol. wt and immunological reactivity as the IgM subunit. The biological significance of the interaction between IgM and leukocyte proteinases is discussed.

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Neurotic disturbances in children can be largely explained by pathogenic attitudes of the parents. Disappointment, guilt and helplessness are neutralised or denied by a multitude of defence mechanisms. The advisory service for parents is concerned with the task to dissolve these defence mechanism and to relieve the disturbed parents-child relations. According to the authors' opinion, parents can arrive at a "child-centred" attitude when the therapist, too, is experienced in his attitude as "child-and-parents-centred". In the group work with parents, a combination of talk groups with "supplementary methods" after Brocher and training groups according to the "defeatless method of conflict overcoming" after Gordon has proved to be suitable. Practical approach and experiences are described.

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Immunohistochemical studies were performed in synovial tissues from 40 patients with rheumatoid arthritis (RA), 9 with juvenile rheumatoid arthritis (JRA), 7 with psoriatic arthritis, and 4 with various rheumatic diseases. Overall synthesis of IgG- and/or IgM-rheumatoid factor (RF) was found in all patients with seropositive RA and JRA, in 75% of patients with seronegative RA, and in 1 patient with psoriatic arthritis. Agglutinator producing cells were found in 77% of the samples from seropositive RA and in 44% and 56% from seronegative RA and JRA patients, respectively. The percentage of IgG plasma cells synthesizing one or more of the 5 types of agglutinators studied was approximately 10% of plasma cells synthesizing IgG-RF. Intercellular and intracellular immune complex deposits were also found in patients with seropositive and seronegative RA and JRA. These findings suggest that synthesis of agglutinators by synovial tissue plasma cells of RA and JRA patients is a distinct--but definitely less prominent--function than that of RF synthesis.

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Using a specific substrate, no leucocyte elastase activity could be detected in 55 synovial fluids, including 29 from patients with rheumatoid arthritis (RA). However, a high percentage of samples contained phagocytic inclusions of elastase, alpha 1-proteinase inhibitor (alpha 1-PI) and alpha 2-macroglobulin (alpha 2-MG) in both the polymorphonuclear (PMN) and mononuclear phagocytes. Immunofluorescence and indirect peroxidase-antiperoxidase staining of articular cartilage (ACA) from 52% of 21 patients with RA and one with juvenile RA (JRA) showed presence of elastase in the superficial layer of microscopically intact but proteoglycan depleted pannus-free ACA. In histologically altered pannus-free RA-ACA superficial elastase deposits were found in 24% of the cases. Adjacent ACA sections contained IgG, C3, alpha 1-PI and rarely alpha 2-MG. RA-ACA below or surrounded by pannus showed close contact with intact and decaying PMN in 62% and 48% of the cases, respectively. ACA specimens from patients with degenerative disease and systemic lupus were negative. These findings strongly suggest that PMN leucocyte elastase is operative in the degradation of RA-ACA and JRA-ACA, and that this activity is largely dependent upon the presence of entrapped immune complexes in such cartilage.

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Human lysosomal elastase cleaves human monoclonal IgG into components that closely resemble the fragments produced by papain digestion. IgG1 produced Fab, Fc and Fab-Fc fragments; cleavage of IgG2 produced F(ab)2, Fab-Fc, Fab and Fc fragments; IgG3 gave rise to almost pure Fab and Fch (Fc covalently joined to the extended hinge region polypeptide of IgG3), and from IgG4, F(ab)2, Fab and Fc fragments were recovered. The relative susceptibilities of the four human IgG subclasses to proteolytic attack by elastase were studied kinetically and showed the following decreasing order of susceptibility: IgG3 > IgG1 > IgG2 > IgG4. The Fab fragment from papain digestion of IgG1 and the corresponding fragment from elastase digestion showed indistinguishable molecular weights and immunochemical identity.

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The synovial fluid (SF) of RA patients contains large amounts of PMN which are well equipped with neutral enzymes to degrade articular cartilage: elastase and cathepsin G, which both destroy proteoglycans and native collagen, as well as 2 types of collagenoases. Indirect evidence suggests that PMN might be important in the destruction of RA articular cartilage. In 19 SF of RA patients no free elastase or collagenase was found. Using immune histochemical methods, we observed that PMN and macrophages of SF contain both elastase and alpha 1-anti-trypsin and alpha 2-macroglobulin. Peripheral PMN - but not monocytes - contain elastase, however both types of cells lack alpha 1-antitrypsin and alpha 2-macroglobulin. Elastase is demonstratable in the superficial layer of pannus free RA articular cartilage. These findings suggest that neutral proteinases from PMN in RA SF are generally neutralized by physiologic inhibitors and removed by phagocytes. The enzyme-inhibitor interaction might be bypassed during "frustrated phagocytosis" so that enzymes like PMN elastase can damage RA articular cartilage.

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The authors report their experience with a special form of psychodrama for the treatment in small groups of neurotic children aged five to fifteen. The acting out of ambivalent, partly unconscious and restrained tendencies and inclinations by playing, i.e., by taking roles in spontaneous performances, is followed by alternation of identification in the group and, possibly, careful indirect verbalization by the therapeutist and/or group, of which the purpose is to arrive at a reorientation of the attitudes of patients and a consolidation of the newly won attitude.

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This article describes the complexity and dynamics of the disease with reference to the diagnosis of "behavioral disorder in the case of normal intelligence". In addition, the authors discuss differences in manifestation and the need for cooperation between experts in pedoneuropsychiatry and clinical psychology, on the one hand, and pedagogics and special education, on the other.

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In those cases where the subjects involved tend to show pathological psychophysical reactions, it is necessary for a cooperative effort to be made by pedagogs, psychologists, and medical experts (pediatricians and pedoneuropsychiatrists). This may be realized by patient-related corrdinative consultations or psychotherapeutic treatment and subsequent individual pedagogic promotion and encouragement depending upon the degree of manifestation and generalization of the disorders. The use of various methods (modified but predominantly pedagogic and directive approaches and chiefly psychological and nondirective methods of treatment, respectively) involve different responsibilities of experts for the process of rehabilitation (in regard to both time and content). Also, the authors show that this is not in contradiction to general objectives of education and training and that every effort is made to reconcile medical and psychological objects with pedagogic requirements.

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