For differences among materials to be easily detected, low variation in in vitro wear tests is desirable. The working hypothesis of this paper was that antagonists standardized for shape and size and according to materials would show mean values similar to those found in natural, non-standardized cusps, and that standardization would lead to a reduction in mean variation. First, the shapes and sizes of palatal cusps of non-erupted human upper third molars were measured. The cusp cupola was best described by the formula y = 0.001 x2 and was symmetrical around the axis of rotation. Up to 200 μm of the y-axis, this parabola corresponded best to a ball radius of 0.6 mm. Based on this information, standardized antagonists were fabricated from both human enamel and steatite. Wear in the occlusal contact area and the wear of opposing conventional ceramic and fine hybrid composite, respectively, were quantified in a computerized chewing simulator. As a control, natural human enamel cusps were used. Standardization of enamel cusps did not reduce the variation of the resulting wear compared with that of non-standardized enamel antagonists. Furthermore, standardization led to significantly different results both in the antagonists and in the opposing restorative materials. Thus, natural enamel antagonists are preferable for the simulation of wear in the occlusal contact area.

Targeted DNA methylation is a technique that aims to methylate cytosines in selected genomic loci. In the most widely used approach a CG-specific DNA methyltransferase (MTase) is fused to a sequence specific DNA binding protein, which binds in the vicinity of the targeted CG site(s). Although the technique has high potential for studying the role of DNA methylation in higher eukaryotes, its usefulness is hampered by insufficient methylation specificity. One of the approaches proposed to suppress methylation at unwanted sites is to use MTase variants with reduced DNA binding affinity. In this work we investigated how methylation specificity of chimeric MTases containing variants of the CG-specific prokaryotic MTase M.SssI fused to zinc finger or dCas9 targeting domains is influenced by mutations affecting catalytic activity and/or DNA binding affinity of the MTase domain. Specificity of targeted DNA methylation was assayed in E. coli harboring a plasmid with the target site. Digestions of the isolated plasmids with methylation sensitive restriction enzymes revealed that specificity of targeted DNA methylation was dependent on the activity but not on the DNA binding affinity of the MTase. These results have implications for the design of strategies of targeted DNA methylation.

Calcitonin and its carboxyl-terminal flanking peptide (PDN-21), also encoded by the calcitonin gene, were measured by RIA in unextracted serum of normal subjects and patients with primary hyperparathyroidism and surgically verified and suspected medullary thyroid carcinoma. Serum PDN-21 was detectable (greater than 0.005 ngeq/ml) in the large majority of normal subjects (92%), and the values increased significantly more in men than women (4.8- and 2.0-fold, respectively; P less than 0.01) in response to 1-min iv calcium injections. Calcitonin was detectable (greater than 0.025 ngeq/ml) in only 25% of normal subjects before iv calcium and became measurable after iv calcium in 88% of men and 41% of women. In patients with chronic hypercalcemia due to primary hyperparathyroidism, PDN-21 and calcitonin were within normal limits. In normal subjects, iv pentagastrin (0.5 microgram/kg BW) did not increase PDN-21, and calcitonin remained undetectable. In 41 medullary thyroid carcinoma patients, basal PDN-21 and calcitonin levels were increased similarly, and they were stimulated in response to iv calcium or iv pentagastrin. In 5 siblings of medullary thyroid carcinoma patients, PDN-21 and calcitonin were increased in response to iv pentagastrin, and we suspect C-cell hyperplasia or medullary thyroid carcinoma. In conclusion, a diagnostically useful RIA for the measurement of PDN-21 in unextracted serum which complements calcitonin measurements has been developed.