Expression of HLA-DR antigens on tumour cells does not contribute to skin reactivity to autologous cholesteryl hemisuccinate (CHS)-treated tumour cells in patients with metastatic melanoma

Summary Skin tests with autologous cholesteryl hemisuccinate (CHS)-treated and untreated cells were performed in ten metastatic melanoma patients. In the majority of cases evident reaction was noted with CHS-treated cells (9/10) while the reaction with untreated cells was mostly negative (7/10). Tumour cell suspensions used for skin tests were characterized for reactivity with monoclonal antibody TAL 1B5 detecting the HLA-DR alpha chain. There were no differences between CHS-treated and untreated cells with respect to HLA-DR expression and no correlation was found between grade of skin reaction to CHS-treated cells and the proportion of HLA-DR positive cells in the injected cell sample.     

Prevention of non-specific interactions of gold-labeled reagents on tissue sections

The protein A-gold technique is amongst the most useful labeling techniques available for light and electron microscopic immunolabeling. Some electron microscopic studies, however, have suggested that protein A-gold, and other protein-gold complexes as well, may bind non-specifically to certain tissue structures, particularly in skin, creating a specious pattern of labeling. We utilized the protein A-gold technique with antiserum to both involucrin and keratin under a variety of conditions to document the specificity of labeling. When the standard conditions were followed, the protein A-gold technique produces highly specific results. These conditions include: 1. the blocking of unreacted aldehyde groups by amination; 2. the blocking of non-specific binding sites on tissue sections by preincubation with inert proteins; and 3. the use of proper concentration of the protein A-gold complex. However, non-specific labeling could be produced if the three components of the standard protocol were omitted. In particular, the use of too concentrated protein A-gold lead to non-specific labeling. We report here also updated working protocols for antigen detection with protein A-gold on semithin Lowicryl K4M and paraffin sections which provide optimal staining results.     

Survival of sheep x goat hybrid inner cell masses after injection into ovine embryos

Modified blastocyst injection techniques were used to inject immunosurgically isolated sheep x goat hybrid inner cell masses (ICM) into ovine blastocysts, with subsequent transfer of composite embryos to ovine recipients. Hybrid embryos were collected from does artificially inseminated with Barbados ram semen. A total of 13 live and 2 aborted offspring resulted from the 34 composite embryos transferred to recipient ewes (38% embryo survival). Of the 15 offspring, 4 exhibited phenotypic hybridism and 2 (13%) of these were determined to be hybrid mean value of -sheep chimeras by karyotype, serum protein and isoenzyme analyses, and fiber identification. Each of the 4 was produced by an injection procedure that involved damage of the ovine host ICM. One additional offspring was unusual in appearance, but the presence of hybrid cells was not proven. Similarly, caprine ICM were immunosurgically isolated and injected into ovine blastocysts that were then transferred to ovine recipients. Of the 13 composite embryos transferred, 12 offspring were produced (92% embryo survival). Eleven were overt goat mean value of -sheep chimeras and, of these, 7 were also blood chimeras. The hybrid ICM was shown to be capable of contributing to normal embryonic and fetal development after injection into an ovine blastocyst but may be less likely to be incorporated with the ovine host ICM than is the caprine ICM.     

Structural and physicochemical requirements of endotoxins for the activation of arachidonic acid metabolism in mouse peritoneal macrophages in vitro

Lipopolysaccharides of different wild-type and mutant gram-negative bacteria, as well as synthetic and bacterial free lipid A, were studied for their ability to activate arachidonic acid metabolism in mouse peritoneal macrophages in vitro. It was found that lipopolysaccharides of deep-rough mutants of Salmonella minnesota and Escherichia coli (Re to Rc chemotypes) stimulated macrophages to release significant amounts of leukotriene C4 (LTC4) and prostaglandin E2 (PGE2). Lipopolysaccharides of wild-type strains (S. abortus equi, S. friedenau) only induced PGE2 and not LTC4 formation. Unexpectedly, free bacterial and synthetic E. coli lipid A were only weak inducers of LTC4 and PGE2 production. Deacylated Re-mutant lipopolysaccharide preparations were inactive. However, co-incubation of macrophages with both deacylated lipopolysaccharide and lipid A lead to the release of significant amounts of LTC4 and PGE2, similar to those obtained with Re-mutant lipopolysaccharide. The significance of the lipid A portion of lipopolysaccharide for the induction of LTC4 was indicated by demonstrating that peritoneal macrophages of endotoxin-low-responder mice or of mice rendered tolerant to endotoxin did not respond with the release of arachidonic acid metabolites on stimulation with Re-mutant lipopolysaccharide and that polymyxin B prevented the Re-lipopolysaccharide-induced LTC4 and PGE2 release. Physical measurements showed that the phase-transition temperatures of both free lipid A and S-form lipopolysaccharide were above 37 degrees C while those of R-mutant lipopolysaccharides were significantly lower (30-35 degrees C). Thus, with the materials investigated, an inverse relationship between the phase-transition temperature and the capacity to elicit LTC4 production was revealed.     

Abnormal centrosomes in cold-treated Drosophila embryos

In this study we examine the effect on the centrosomes of cold treatment of early Drosophila embryos. Prolonged cold treatment during the mitotic divisions which lead to the formation of the blastoderm causes arrest at metaphase of the nuclear divisions. When examined with immunofluorescence microscopy the mitotic spindles show marked pole splitting with the formation of supernumerary and irregularly sized centers, all able to nucleate microtubules. In embryos recovered for longer periods the additional organizing centers become ring-shaped and lose their nucleating properties. Cold treatment of embryos during the cellularization of the blastoderm results in marked fragmentation of the centrosomes, but nucleating capacity is preserved. Sometimes the centrioles come away from the pericentriolar material and their structure is seen to be modified.     

DNA ligase and the pyridine nucleotide cycle in Salmonella typhimurium.

Bacterial DNA ligases use NAD as an energy source. In this study we addressed two questions about these enzymes. First, what is the physiological consequence of completely removing the NAD-dependent enzyme and replacing it with an ATP-dependent DNA ligase? We constructed Salmonella typhimurium strains in which the endogenous NAD-dependent DNA ligase activity was inactivated by an insertion mutation and the ATP-dependent enzyme from bacteriophage T4 was provided by a cloned phage gene. Such strains were physiologically indistinguishable from the wild type, even under conditions of UV irradiation or treatment with alkylating agents. These results suggest that specific functional interactions between DNA ligase and other replication and repair enzymes may be unimportant under the conditions tested. Second, the importance of DNA ligation as the initiating event of the bacterial pyridine nucleotide cycle was critically assessed in these mutant strains. Surprisingly, our results indicate that DNA ligation makes a minimal contribution to the pyridine nucleotide cycle; the Salmonella strains with only an ATP-dependent ligase had the same NAD turnover rates as the wild-type strain with an NAD-dependent ligase. However, we found that NAD turnover was significantly decreased under anaerobic conditions. We suggest that most intracellular pyridine nucleotide breakdown occurs in a process that protects the cell against oxygen damage but involves a biochemical mechanism other than DNA ligation.     

Modeling rendille household herd composition

Previous analysis of Rendille household herd composition revealed a transition from camel to cattle ownership for sedentary impoverished Rendille pastoralists of northern Kenya. In an attempt to delineate determinants of livestock holdings, logistic regression analysis of 112 household herds from the Rendille settlement of Korr, Marsabit District, Kenya was undertaken. Results indicated that household wealth, measured in present livestock holdings, past drought losses, and livestock sales, formed better predictors of cattle ownership than did household characteristics pertaining to labor supply, wage earners, age-set membership, and birth order of household head. These results are discussed in light of pastoral strategies designed to minimize risk.     

Drought and economic differentiation among Ariaal pastoralists of Kenya

This paper examines the effects of the 1984 drought upon household wealth differences in a community of Ariaal pastoralists of northern Kenya. The database consists of 1985 post-drought livestock counts and informants' statements of species-specific drought loss, compared to 1976 livestock counts on the same 38 households. The analysis confirms the hypothesis that the drought resulted in increased household wealth inequalities. It is suggested that the combination of differential herd growth, differential participation in the cash market, and differential loss to the drought has contributed to a polarization within Ariaal of rich and poor, resulting in rural proletarianization and urban migration.     

Ultrastructural immuno-localization of tropoelastin in the chick eye

Summary Immuno-gold labeling at the electron-microscopy level was used to investigate the distribution of tropoelastin in the chick eye. Intense staining was found in the amorphous part of mature elastic fibers in different regions of the organ. In elaunin fibers, both the amorphous core and the surrounding microfibrils were clearly labeled. In addition, reactive sites were detected in the oxitalan fibers of the stroma of the cornea and in Descemet's membrane, which showed a gradient of reactive sites increasing from the center toward the periphery. Oxitalan fibers of the stroma often fused with Descemet's membrane; the pattern of immunological staining suggested a continuity between the two structures. In the ciliary zonule, labeling for tropoelastin was observed in discrete areas on the bundles of microfibrils. The results show a complex structural organization of elastic tissue; this may be important in endowing the various parts of the eye with different mechanical properties.