Single-locus control of the mast cell population in mouse skin

The number of mast cells in connective tissue from dorsal skin varied markedly among mouse strains. Inbred strains of mice were typed into three groups, high (NC and NZB mice), low (B6, B10, and BALB/c mice), and intermediate (C3H/He and DBA/2 mice), by their mast cell content in the skin. However, the strain differences in the number of mast cells was marginal at the age of weaning but became distinct with age. This could be explained mainly by the frequently observed clustering of mast cells in adult NC and NZB mice and the rarely observed clustering in younger mice as well as in adult B10 and BALB/c mice. The breeding experiment revealed that the difference in the number of mast cells between NC and B10 mice was controlled by a single autosomal dominant locus, for which we propose the designation Mcr (mast cell regulator). The role of the Mcr locus with regard to the frequency of the mast cell population in connective tissue is discussed.     

The limited proteolysis of tumor necrosis factor-α

The limited proteolysis of human recombinant TNF- by trypsin yields two stable products resulting from cleavage after Arg6 and Arg44. In solution these two products remain associated together in a trimer with a Stokes' radius slightly greater than the radius of intact TNF- and, therefore, could not be separated from each other under nondenaturing conditions. This limited digest retains at least 20% of the activity of the original TNF- sample, and has a tertiary structure that is similar to that of the native protein by circular dichroism. On the other hand, incorrectly folded, inactive TNF- undergoes extensive digestion following similar treatment with trypsin. These results indicate that the active form of TNF- has a tight core structure which is maintained afterN-terminal cleavage and removal.     

Suppressor cells in the effector phase of autologous cytotoxic reactions in cancer patients

Summary Cytotoxicity was induced in lymphocytes (CL) from 10 out of 15 patients by autologous mixed lymphocyte tumor cell culture and further cultivation with recombinant interleukin-2. In cells from 3 of the 10 patients, cytotoxicity was suppressed by more than 50% when autologous peripheral blood mononuclear cells (PBMC) from the patients with large tumors were added to the autologous killing system. The cells responsible for suppressing the cytotoxicity in the effector phase were adherent or nonadherent to plastic depending on the patient examined. The T cell fraction from 1 patient significantly suppressed the cytotoxic activity, and this suppression was seen only in the autologous system. On the other hand, plastic adherent cells but not T cells from PBMC of 2 subjects suppressed the cytotoxic activity of CL. The reason why the main cell population suppressing the CL activity differed among the patients is unclear. However, the findings that the suppression was mostly abrogated following resection of the tumor mass suggested that suppressor cells, either of macrophage lineage or T cells, are induced in patients with a large tumor mass. This speculation is supported by the finding that the PBMC from a patient with tumor recurrence regained the suppressive activity.     

Effects of chemically defined tannins on poly (ADP-ribose) glycohydrolase activity.

Three classes of chemically defined tannins, gallotannins, ellagitannins and condensed tannins were examined for their inhibitory activities against purified poly (ADP-ribose) glycohydrolase. Ellagitannins showed higher inhibitory activities than gallotannins. In contrast, condensed tannins, which consist of an epicathechin gallate (ECG) oligomer without a glucose core were not appreciably inhibitory. Kinetic analysis revealed that the inhibition of ellagitannins was competitive with respect to the substrate poly(ADP-ribose), whereas gallotannins exhibited mixed-type inhibition. These results suggest that conjugation with glucose of hexahydroxy-diphenoyl (HHDP) group, which is a unique component of ellagitannins, potentiated the inhibitory activity, and that the structure of ellagitannins may have a functional domain which competes with poly(ADP-ribose) on the poly(ADP-ribose) glycohydrolase molecule.     

Killer cells induced by stimulation with allogeneic tumor cells and subsequent culture with recombinant interleukin-2

Summary Peripheral blood lymphocytes were cultured for 5 days with allogeneic tumor cells (allogeneic mixed lymphocyte/tumor cell culture), and subsequently cultured with recombinant interleukin-2 for 12 days. These cultured cells were found to be cytotoxic to autologous tumor cells. Results of two-color analysis using monoclonal antibodies to cell markers showed that more than 80% of their cultured cells were CD3⁺ cells, and CD4⁺ cells showed a higher distribution than CD8⁺ cells. However, CD8⁺ cells had a much higher killing activity with autologous tumor than did CD4⁺ cells, when estimated by an elimination study using monoclonal antibodies to T cell phenotypes and complement. The cold-target inhibition test showed that the cytotoxicity of these cells for autologous tumor cells was inhibited by unlabeled autologous tumor cells but not by unlabeled stimulator cells. Furthermore, about 40% of the cytotoxicity was suppressed by blocking of HLA class I antigen with a monoclonal antibody on autologous tumor cells. Thus, cytotoxic activity of lymphocytes to autologous tumor restricted by target cell HLA class I antigen is possibly induced by allogeneic tumor-stimulation.     

Analysis of bacterial populations in a grassland soil according to rates of development on solid media

Abstract The process of colony formation by bacteria from grassland soil sampled in April, July and September was simulated by a colony-forming curve (CFC). The CFC was a super-imposition of several component curves (cCFC) given theoretically by the first order reaction (FOR) model [3,6]. The pattern of FOR model curves was not influenced by the time of sampling and four cCFCs were always recognized during an incubation period of 160 h. It was considered that the CFC describes an inherent property of the bacterial population of the field. Bacterial isolates were obtained from colonies produced in each of four cCFCs on agar plates. Isolates corresponding to one cCFC were classified as one group. The bacterial isolates were characterized by morphological and physiological tests and subsequently clustered. Few oligotrophic bacteria were obtained among bacteria which produced visible colonies within 63 h of incubation time. On the other hand, approx. 50% of bacteria which produced v colonies after 63 h were oligotrophic bacteria. The time required for the appearance of the first colony, t _r of the FOR model, was very similar in the isolates belonging to one group. A close linear relationship was observed between t _r value and doubling time of isolates.     

Membrane potential in liposomes measured by the transmembrane distribution of 86Rb+, tetraphenylphosphonium or triphenylmethylphosphonium: effect of cholesterol in the lipid bilayer

Valinomycin-induced potassium diffusion potential (delta psi, inside negative) in the liposomes made of phosphatidylcholine and various amounts of cholesterol was measured by uptake of 86Rb+, tetraphenylphosphonium (TPP+) or triphenylmethylphosphonium (TPMP+). In any liposome, the values of membrane potential obtained by 86Rb+ uptake (delta psi Rb) agreed well with those calculated from the imposed potassium concentration gradient using the Nernst equation, and were not affected by the presence of cholesterol. However, both delta psi TPP and delta psi TPMP showed smaller values than delta psi Rb when the cholesterol content in liposomes increased. delta psi TPMP at a stationary state was much smaller than delta psi TPP. The orientational order parameter of the lipids' bilayer with various cholesterol content was estimated from fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene. The results indicated that the permeation of TPP+ or TPMP+ into liposomes containing a large amount of cholesterol is strongly restricted by the high ordering of phosphatidylcholine acyl chains.     

Actin-fragmin interactions as revealed by chemical cross-linking

A one to one complex of actin and fragmin (a capping protein from Physarum polycephalum plasmodia) was cross-linked with 1-ethyl-3-[3-(dimethylamino)propyl] carbodiimide. The cross-linking reaction generated two cross-linked products with slightly different molecular weights (88 000 and 90 000) as major species. They were cross-linked products of one actin and one fragmin. The cross-linking site of fragmin in the actin sequence was determined by peptide mappings [Sutoh, K. (1982) Biochemistry 21, 3654-3661] after partial chemical cleavages of cross-linked products with hydroxylamine. The results indicated that the N-terminal segment of actin spanning residues 1-12 participated in cross-linking with fragmin. The cross-linker used in this study covalently bridges lysine side chains and side chains of acidic residues when they are in direct contact. Therefore, it seems that acidic residues in the N-terminal segment of actin (Asp-1, Glu-2, Asp-3, Glu-4, and Asp-11), at least some of them, are in the binding site of fragmin. It has already been shown that the same acidic segment of actin is in the binding site of myosin or depactin (an actin-depolymerizing protein isolated from starfish oocytes). We suggest that the unusual amino acid sequence of the N-terminal segment of actin makes its N-terminal region a favorable anchoring site for various types of actin-binding proteins.