Isolation and characterization of a lectin from tulip bulbs, Tulipa gesneriana

A lectin, which agglutinated specifically the yeast cells of the Saccharomyces genus, was isolated from tulip bulbs (Tulipa gesneriana) using affinity chromatography on mannan-Sepharose 4B. Its relative molecular mass was determined by gel filtration to be approximately 67,000. On polyacrylamide gel electrophoresis in sodium dodecyl sulfate, a relative molecular mass of 17,000 was obtained, suggesting that the lectin is a tetramer. Binding studies performed with iodinated lectin indicated that Saccharomyces cerevisiae cells contained approximately 5.7 X 10(6) binding sites per cell, whereas little binding was observed with yeasts other than the Saccharomyces genus, bacteria and animal erythrocytes. D-Mannose, D-mannose 6-phosphate, L-fucose and L-fucosylamine were potent inhibitors of the lectin binding to S. cerevisiae cells, while, D-glucose, D-galactose and D-mannosamine were inactive, indicating that hydroxyl group at C-2 of D-mannose was essential for the lectin binding. Furthermore, inhibition experiments, using various manno-oligosaccharides, suggested that the lectin recognized (1----6)-linked manno-oligosaccharide units larger than mannobiose.     

The role of adherent accessory cells in the generation of effector suppressor T cells

The role of accessory cell populations in the generation of effector suppressor (Ts3) cells was studied. By using an in vitro culture system, it was previously determined that the induction of NP-specific effector suppressor activity requires T cells, antigen, and an anti-idiotypic B cell population. We now demonstrate that the generation of Ts3 cells in this system also requires accessory cells. The accessory population appears to play a role in the processing and presentation of antigen. These antigen-presenting accessory cells are required early in the induction phase of Ts3 generation. These accessory cells can present NP coupled to immunogenic or non-immunogenic polypeptide carriers, including polymers of L-amino acids. However, NP coupled to polymers of poorly metabolized D-amino acids fail to induce suppressor T cell generation. Furthermore, the data demonstrate that an H-2 homology must exist between the Ts3 precursors and the antigen-presenting cell population if suppressor activity is to be generated. We also characterize the differential genetic restrictions that govern the induction of Ts3 cells that control suppression of either T cell or B cell responses. The data suggest that although I-J region encoded gene products control the induction and effector phases of suppressor cell activity as measured on T cell responses, the suppression of B cell responses appear to be controlled by I-A gene products. Possible cellular mechanisms that might explain these findings are discussed.     

Alloantigen presentation by B cells: two types of alloreactive T cell hybridomas, B cell-reactive and B cell-nonreactive

T cell-depleted, Sephadex G-10-passed unstimulated splenic B cells from C57BL/6 mice stimulated splenic T cells from CKB mice to produce IL 2 and to proliferate. The stimulatory ability of the unstimulated B cells was eliminated by 4000 rad irradiation of the unstimulated stimulator B cells. LPS-activated B cells could stimulate responder T cells more efficiently than unstimulated B cells. For further analysis of allostimulation by B cells, we established a series of alloreactive T cell hybridomas. Forty-five percent of these alloreactive T cell hybridomas could be stimulated to produce IL 2 by either macrophage-dendritic cells or unstimulated B cells. Fifty-five percent of these alloreactive T cell hybridomas could be stimulated by macrophage-dendritic cells but not by unstimulated B cells. T cell hybridomas that were not reactive with unstimulated B cells were also nonreactive to LPS-activated B cells. Analysis of two representative I-Ab-reactive T cell hybridoma clones, B cell-reactive clone CB-11.4 and B cell-nonreactive clone HTB-9.3, revealed again that the stimulatory ability of unstimulated B cells was sensitive to 4000 rad irradiation in the activation of CB-11.4 clone and that CB-11.4 could be stimulated more efficiently by LPS-activated B cells than by unstimulated B cells, but HTB-9.3 could not be stimulated by LPS-activated B cells. Thus, there may be two distinct types of T cells in the alloreaction: B-cell-reactive and B cell-nonreactive.     

A genetically restricted suppressor factor that requires interaction with two distinct targets

We have previously described a genetically restricted suppressor factor (TsF3) that suppresses the terminal phases of the contact sensitivity response. The activity of TsF3 is restricted by genes in the H-2 (I-J) and Igh complexes. This report analyzes the mechanisms responsible for these genetic restrictions. One cellular target of TsF3 is an I-J-bearing antigen-presenting cell population that is sensitive to low doses of cyclophosphamide. To elicit suppression I-J homology is required between this antigen-presenting cell population and the TsF3 donor. In contrast, the Igh-linked genetic restriction exists between TsF3 and an unprimed cell population present in the recipient. These findings suggest that under these experimental conditions TsF3 acts by bridging the APC with cells of the host. Finally, we demonstrated that nonspecific bystander or cognate suppression can be mediated by TsF3, provided specific antigen is present in the site of the ongoing T cell response.     

Role of lymphokines in regulation of macrophage differentiation

The regulatory mechanism of guinea pig lymphokines was investigated in regard to differentiation of myeloid cells to macrophages. The Ml-cell line, established from a myeloid leukemia of an SL-strain mouse, was induced to differentiate in vitro into mature macrophages possessing Fc receptors and the ability to phagocytize latex particles by treatment with crude lymphokines. Both concanavalin A- and antigen-induced lymphokines showed the differentiation-inducing factor (D factor) activity. However, macrophage migration inhibitory factor/ macrophage activation factor (MIF/MAF) purified by an immunoadsorbent column with anti-MIF antibody had no such an activity. The D-factor activity was detected in the lymphokine preparation that was not retained on the immunoadsorbent column. In contrast, colony-stimulating factor (CSF) was adsorbed to the immunoadsorbent column, and could be recovered in the purified MIF/MAF preparation. These findings suggest that the molecular entity of D factor is distinct from MIF/ MAF and CSF. A culture supernatant of guinea pig peritoneal macrophages activated with MIF/ MAF (CSF) exhibited strong D-factor activity. However, the supernatant possessed rather reduced CSF activity as compared to that of the original MIF/MAF (CSF) preparation. Thus, MIF/MAF may play an important role in macrophage differentiation by regulating the production of D factor or CSF from macrophages.     

Location in Muscarinic Acetylcholine Receptors of Sites for [3H]Propylbenzilcholine Mustard Binding and for Phosphorylation with Protein Kinase C

Muscarinic acetylcholine receptors purified from porcine cerebra or atria were covalently labeled with [3H]propylbenzilylcholine mustard ([3H]PrBCM), and then the labeled receptors were subjected to limited hydrolysis with trypsin, V8 protease, and lysyl endopeptidase, followed by analysis involving sodium dodecyl sulfate-polyacrylamide gel electrophoresis, fluorography, autoradiography, or immunostaining. The labeled peptides were located on the basis of their reactivity with antibodies raised against three synthetic peptides with partial sequences of the m1 or m2 receptor, and of their sensitivity to endoglycosidase F, which was taken as evidence that they contain glycosylation sites near the N terminus. The [3H]PrBCM-binding site in both cerebral and atrial receptors was found to be located between the N terminus and the second intracellular loop, because the size of the smallest deglycosylated peptide that contained both the [3H]PrBCM-binding and glycosylation sites was approximately 16 kDa. Cerebral receptors were 32P-phosphorylated with protein kinase C, and the major phosphorylation sites in cerebral muscarinic receptors were found to be located in a C-terminal segment including a part of the third intracellular loop, because a 32P-labeled peptide of 12-14 kDa reacted with anti-(m1 C-terminal peptide) antiserum. The presence of an intramolecular disulfide bond, probably between Cys 98 and Cys 178 in the first and second extracellular loops, respectively, was suggested by the finding that a peptide of approximately 17 kDa containing the [3H]PrBCM-binding site, but not the glycosylation sites, was partly converted to a peptide of approximately 12 kDa on treatment with beta-mercaptoethanol.     

Leukotriene A4 hydrolase from guinea pig lung: the presence of two catalytically active forms

Leukotriene A4 hydrolase was purified to apparent homogeneity from the guinea pig lung. The molecular weight was determined to be 70 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme exhibited two active forms with different pI values (5.7 and 5.4) depending on the presence or absence of SH-reducing reagents during purification procedures. No significant differences were observed between both forms of the enzyme as regards the catalytic properties. The N-terminal 20 amino acid sequence (PEVVDTXSLASPATVXRTKH) showed a 90% identity to the human enzyme with a constitutive substitution of Ile-3 and Ser-14 (human) by Val-3 and Thr-14 (guinea pig), respectively.     

Mother-infant interactions of wild-born,individually-caged cynomolgus monkeys (Macaca fascicularis) during the first 14 weeks of infant life

This study documents age-related changes in the interactions of wild-born cynomolgus macaque mothers and their infants living in individual cages during the first 14 weeks of infant life. Body contact between mother and infant, maternal holding, and infant sucking were found to decrease, and the mothers showed an increased frequency of aggression toward their infants with age. These results were broadly similar to those reported for mother-infant interactions in other macaques living in social groups. Nevertheless, a clear difference between the present cynomolgus macaques and other macaques in social groups was apparent. The cynomolgus macaque mothers tended to permit their infants to move about freely without displaying maternal protectiveness such as restraint or retrieval, unlike other macaque mothers in social groups. Such maternal behaviors might derive from the experience of living in individual cages for many years and the relative safety of living in individual cages. The lack of maternal restraint and retrieval could be responsible for the observed sex differences in behavior: male infants moved more actively, and broke, and made contact with their mothers more frequently than did female infants. Moreover, mothers of female infants held and groomed them more frequently and were less aggressive toward them.