Isolation of a protease inhibitor from tissues resistant to tumor invasion

We have purified to homogeneity the major trypsin inhibitors from both bovine cartilage and aorta, two tissues reported to be highly resistant to invasion. The two inhibitors appear to be identical and they resemble the Kunitz inhibitor with respect to molecular weight, amino acid composition, range of susceptible proteases, and antigenicity. Each of these inhibitors accounts for 100% of the antitrypsin activity found in extracts of bovine cartilage and aorta.     

Movements and associations of ribosomal subunits in a secretory cell during growth inhibition by starvation

In Chironomus tentans salivary gland cells, the cytoplasm can be dissected into concentric zones situated at increasing distances from the nuclear envelope. After RNA labeling, the newly made ribosomal subunits are found in the cytoplasm mainly in the neighborhood of the nucleus with a gradient of increasing abundance towards the periphery of the cell. The gradient for the small subunit lasts for a few hours and disappears entirely after treatment with puromycin. The large subunit also forms a gradient but one which is only partially abolished by puromycin. The residual gradient which which is resistant to the addition of the drug is probably due to the binding of some large ribosomal units to the membranes of the endoplasmic reticulum (J.-E. Edstrom and u. Lonn. 1976. J. Cell Biol. 70:562-572, and U. Lonn and J.-E. Edstrom. 1976. J. Cell. Biol. 70:573-580). If growth is inhibited by starvation, only the puromycin-sensitive type gradient is observed for the large subunit, suggesting that the attachment of these newly made subunits to the endoplasmic reticulum membranes will not occur. If, on the other hand, the drug-resistant gradient is allowed to form in feeding animals, it is conserved during a subsequent starvation for longer periods than in control feeding animals. This observation provides a further support for an effect of starvation on the normal turnover of the large subunits associated with the endoplasmic reticulum. These results also indicate a considerable structural stability in the cytoplasm of these cells worth little or no gross redistribution of cytoplasmic structures over a period of at least 6 days.     

Effects of protein perturbants on phospholipid bilayers

Series of alcohols, amides, ureas, and sulfoxides with increasingly longer hydrocarbon chains have been shown to lower progressively the thermal denaturation temperature of proteins. This effect is presumably due to a hydrophobic interaction between the solute and nonpolar domains of the protein. Theoretically, these interactions should occur between the solute and any macromolecular structure having a nonpolar region to which the solute has access. A recent review by Arakawa et al. has summarized evidence for such an interaction between organic solutes and proteins and suggested that these interactions are favored at higher temperatures. The present study investigates the effects of several classes of compounds on the stability of phospholipid vesicles. The results show that many compounds that are known to perturb protein function also destabilize phospholipid bilayers as reflected by solute-induced loss of vesicle contents.     

A method for the interpretation of signals in afferent nerves from muscle spindles

A method is proposed for the interpretation of the signals in sensory nerve fibres. It is applicable to systems in which the transfer function between the input to a sense organ and the action potential firing frequency is known. In the present case, the chelonian muscle spindle is considered since its output to ramphold-relase and sinusoidal stretches can be rather accurately simulated.     

Rapid Copper Acquisition by Developing Murine Mesothelioma: Decreasing Bioavailable Copper Slows Tumor Growth,Normalizes Vessels and Promotes T Cell Infiltration

Copper, an essential trace element acquired through nutrition, is an important co-factor for pro-angiogenic factors including vascular endothelial growth factor (VEGF). Decreasing bioavailable copper has been used as an anti-angiogenic and anti-cancer strategy with promising results. However, the role of copper and its potential as a therapy in mesothelioma is not yet well understood. Therefore, we monitored copper levels in progressing murine mesothelioma tumors and analyzed the effects of lowering bioavailable copper. Copper levels in tumors and organs were assayed using atomic absorption spectrophotometry. Mesothelioma tumors rapidly sequestered copper at early stages of development, the copper was then dispersed throughout growing tumor tissues. These data imply that copper uptake may play an important role in early tumor development. Lowering bioavailable copper using the copper chelators, penicillamine, trientine or tetrathiomolybdate, slowed in vivo mesothelioma growth but did not provide any cures similar to using cisplatin chemotherapy or anti-VEGF receptor antibody therapy. The impact of copper lowering on tumor blood vessels and tumor infiltrating T cells was measured using flow cytometry and confocal microscopy. Copper lowering was associated with reduced tumor vessel diameter, reduced endothelial cell proliferation (reduced Ki67 expression) and lower surface ICAM/CD54 expression implying reduced endothelial cell activation, in a process similar to endothelial normalization. Copper lowering was also associated with a CD4⁺ T cell infiltrate. In conclusion, these data suggest copper lowering is a potentially useful anti-mesothelioma treatment strategy that slows tumor growth to provide a window of opportunity for inclusion of other treatment modalities to improve patient outcomes.     

The effect of the elongation of the proximal aorta on the estimation of the aortic wall distensibility

The compliance of the proximal aortic wall is a major determinant of cardiac afterload. Aortic compliance is often estimated based on cross-sectional area changes over the pulse pressure, under the assumption of a negligible longitudinal stretch during the pulse. However, the proximal aorta is subjected to significant axial stretch during cardiac contraction. In the present study, we sought to evaluate the importance of axial stretch on compliance estimation by undertaking both an in silico and an in vivo approach. In the computational analysis, we developed a 3-D finite element model of the proximal aorta and investigated the discrepancy between the actual wall compliance to the value estimated after neglecting the longitudinal stretch of the aorta. A parameter sensitivity analysis was further conducted to show how increased material stiffness and increased aortic root motion might amplify the estimation errors (discrepancies between actual and estimated distensibility ranging from − 20 to − 62%). Axial and circumferential aortic deformation during ventricular contraction was also evaluated in vivo based on MR images of the aorta of 3 healthy young volunteers. The in vivo results were in good qualitative agreement with the computational analysis (underestimation errors ranging from − 26 to − 44%, with increased errors reflecting higher aortic root displacement). Both the in silico and in vivo findings suggest that neglecting the longitudinal strain during contraction might lead to severe underestimation of local aortic compliance, particularly in the case of women who tend to have higher aortic root motion or in subjects with stiff aortas.

     

Human Germline Antibody Gene Segments Encode Polyspecific Antibodies

Structural flexibility in germline gene-encoded antibodies allows promiscuous binding to diverse antigens. The binding affinity and specificity for a particular epitope typically increase as antibody genes acquire somatic mutations in antigen-stimulated B cells. In this work, we investigated whether germline gene-encoded antibodies are optimal for polyspecificity by determining the basis for recognition of diverse antigens by antibodies encoded by three V_H gene segments. Panels of somatically mutated antibodies encoded by a common V_H gene, but each binding to a different antigen, were computationally redesigned to predict antibodies that could engage multiple antigens at once. The Rosetta multi-state design process predicted antibody sequences for the entire heavy chain variable region, including framework, CDR1, and CDR2 mutations. The predicted sequences matched the germline gene sequences to a remarkable degree, revealing by computational design the residues that are predicted to enable polyspecificity, i.e., binding of many unrelated antigens with a common sequence. The process thereby reverses antibody maturation in silico. In contrast, when designing antibodies to bind a single antigen, a sequence similar to that of the mature antibody sequence was returned, mimicking natural antibody maturation in silico. We demonstrated that the Rosetta computational design algorithm captures important aspects of antibody/antigen recognition. While the hypervariable region CDR3 often mediates much of the specificity of mature antibodies, we identified key positions in the V_H gene encoding CDR1, CDR2, and the immunoglobulin framework that are critical contributors for polyspecificity in germline antibodies. Computational design of antibodies capable of binding multiple antigens may allow the rational design of antibodies that retain polyspecificity for diverse epitope binding.     

Ecology and genetics of hybrid zones in the southern African Pycnonotus bulbul species complex

Lloyd, P., Craig, A.J.F.K., Hulley, P.E., Essop, M.F., Bloomer, P. & Crowe, T.M. 1997. Ecology and genetics of hybrid zones in the southern African Pycnonotus bulbul species complex. Ostrich 68 (2–4): 90–96.

The closely related Blackeyed Bulbul Pycnonotus barbatus, Cape Bulbul P. capensis and Redeyed Bulbul P. nigricans have parapatric to locally sympatric distributions within southern Africa. Extensive hybridization along narrow transition zones between each of the three species pairs is described in a region of the Eastern Cape province, South Africa. The transition zones coincide with ecotones between different vegetation types, which in turn follow escarpments or mountain ranges. The lack of population density depressions within the hybrid zones, together with the variability of the hybrids, suggests the hybrids are viable. Sharp step clines in various phenotypic characters are described across the P. barbatus/P. nigricans hybrid zone. A mtDNA analysis found evidence of possible introgression between P. barbatus and P. capensis. All eight P. barbatus x P. nigricans hybrids analysed possessed P. barbatus mtDNA, suggesting the existence of either positive assortative mating or strong directional selection, but our data are unable to distinguish which. Our results do not support the dynamic-equilibrium model, but are compatible with the bounded-hybrid-superiority model. We conclude that the maintenance of the parapatric distributions of the different taxa is due mainly to differences in environmentally-associated fitness between parental phenotypes or among parental and hybrid phenotypes along an ecotone, with the narrowness of the hybrid zones maintained by the steepness of the environmental gradients crossing them.     

A preliminary assessment of congruence between biodiversity patterns in Afrotropical forest birds and forest mammals

Burgess, N., de Klerk, H., Fjeldsá, J., Crowe, T. & Rahbek, R. 2000. A preliminary assessment of congruence between biodiversity patterns in Afrotropical forest birds and forest mammals. Ostrich 71 (1 & 2): 286–291.

Databases compiled for forest birds and forest mammals in the Afrotropics were tested for congruence of overall patterns and hotspots of species richness and endemism. We also looked at how well a near-minimum set of priority areas for one taxon catered for the second taxon. Overall species richness and richness hotspots of forest birds were significantly correlated with those of forest mammals, as was the case for overall endemism. Endemism hotspots for forest birds and mammals were not significantly correlated. The near-minimum set for forest birds represented 136 (76.5%) forest mammal species. The near-minimum set for forest mammals represented 350 (93.62%) forest bird species. However, to represent all forest mammals three times each, 51 grids were needed in addition to the 78 chosen as a near-minimum set for forest birds, and to represent all forest birds three times each, 43 more grids were needed in addition to the 80 selected for forest mammals. There is some congruence between the patterns of richness, endemism and near-minimum sets for forest birds and mammals in the Afrotropics, but the one taxon does not provide the ideal conservation solution for the other. Further refinement of the databases used in this paper would allow for more rigorous testing of congruence between these two groups.