Evaluation of demineralised,freeze-dried,irradiated bone allografts in the treatment of osseous defects in the oral cavity

Demineralised, freeze-dried bone allografts (DFDBA) have been used extensively by dentists in the treatment of periodontal and periapical osseous defects resulting from inflammatory diseases. Their use in India however, is limited by the availability of quality allografts and the high cost of imported alternatives. A study was conducted to assess the osteogenic potential of DFDBA prepared for the first time in India by the Tata Memorial Hospital (TMH) Tissue Bank. The DFDBA was used in the treatment of osseous defects after removal of periapical lesions associated with devitalised teeth in 10 healthy patients. At the 6-month recall visit all the patients showed a remarkable decrease in the grades of mobility, and 9 out of the 10 patients showed radiographic evidence of complete healing of the osseous defects with evidence of normal bony trabaeculae. These findings indicate that the indigenously prepared DFDBA is a cost effective, biocompatible material with osteogenic potential that can be used effectively in treating osseous defects of periapical lesions associated with non vital teeth.     

“Missing perikymata”—fact or fiction? A study on chimpanzee (Pan troglodytes verus) canines

Recently, a lower than expected number of perikymata between repetitive furrow‐type hypoplastic defects has been reported in chimpanzee canines from the Fongoli site, Senegal (Skinner and Pruetz: Am J Phys Anthropol 149 (2012) 468–482). Based on an observation in a localized enamel fracture surface of a canine of a chimpanzee from the Taï Forest (Ivory Coast), these authors inferred that a nonemergence of striae of Retzius could be the cause for the “missing perikymata” phenomenon in the Fongoli chimpanzees. To check this inference, we analyzed the structure of outer enamel in three chimpanzee canines. The teeth were studied using light‐microscopic and scanning‐electron microscopic techniques. Our analysis of the specimen upon which Skinner and Pruetz (Am J Phys Anthropol 149 (2012) 468–482) had made their original observation does not support their hypothesis. We demonstrate that the enamel morphology described by them is not caused by a nonemergence of striae of Retzius but can be attributed to structural variations in outer enamel that result in a differential fracture behavior. Although rejecting the presumed existence of nonemergent striae of Retzius, our study provided evidence that, in furrow‐type hypoplastic defects, a pronounced tapering of Retzius increments can occur, with the striae of Retzius forming acute angles with the outer enamel surface. We suggest that in such cases the outcrop of some striae of Retzius is essentially unobservable at the enamel surface, causing too low perikymata counts. The pronounced tapering of Retzius increments in outer enamel presumably reflects a mild to moderate disturbance of the function of late secretory ameloblasts. Am J Phys Anthropol 157:276–283, 2015. © 2015 Wiley Periodicals, Inc.     

Assembly defects induce oxidative stress in inherited mitochondrial complex I deficiency

Complex I (CI) deficiency is the most common respiratory chain defect representing more than 30% of mitochondrial diseases. CI is an L-shaped multi-subunit complex with a peripheral arm protruding into the mitochondrial matrix and a membrane arm. CI sequentially assembled into main assembly intermediates: the P (pumping), Q (Quinone) and N (NADH dehydrogenase) modules. In this study, we analyzed 11 fibroblast cell lines derived from patients with inherited CI deficiency resulting from mutations in the nuclear or mitochondrial DNA and impacting these different modules. In patient cells carrying a mutation located in the matrix arm of CI, blue native-polyacrylamide gel electrophoresis (BN-PAGE) revealed a significant reduction of fully assembled CI enzyme and an accumulation of intermediates of the N module. In these cell lines with an assembly defect, NADH dehydrogenase activity was partly functional, even though CI was not fully assembled. We further demonstrated that this functional N module was responsible for ROS production through the reduced flavin mononucleotide. Due to the assembly defect, the FMN site was not re-oxidized leading to a significant oxidative stress in cell lines with an assembly defect. These findings not only highlight the relationship between CI assembly and oxidative stress, but also show the suitability of BN-PAGE analysis in evaluating the consequences of CI dysfunction. Moreover, these data suggest that the use of antioxidants may be particularly relevant for patients displaying a CI assembly defect.     

Synthetic symmetrization in the crystallization and structure determination of CelA from Thermotoga maritima

Protein crystallization continues to be a major bottleneck in X‐ray crystallography. Previous studies suggest that symmetric proteins, such as homodimers, might crystallize more readily than monomeric proteins or asymmetric complexes. Proteins that are naturally monomeric can be made homodimeric artificially. Our approach is to create homodimeric proteins by introducing single cysteines into the protein of interest, which are then oxidized to form a disulfide bond between the two monomers. By introducing the single cysteine at different sequence positions, one can produce a variety of synthetically dimerized versions of a protein, with each construct expected to exhibit its own crystallization behavior. In earlier work, we demonstrated the potential utility of the approach using T4 lysozyme as a model system. Here we report the successful application of the method to Thermotoga maritima CelA, a thermophilic endoglucanase enzyme with low sequence identity to proteins with structures previously reported in the Protein Data Bank. This protein had resisted crystallization in its natural monomeric form, despite a broad survey of crystallization conditions. The synthetic dimerization of the CelA mutant D188C yielded well‐diffracting crystals with molecules in a packing arrangement that would not have occurred with native, monomeric CelA. A 2.4 Å crystal structure was determined by single anomalous dispersion using a seleno‐methionine derivatized protein. The results support the notion that synthetic symmetrization can be a useful approach for enlarging the search space for crystallizing monomeric proteins or asymmetric complexes.     

Co-variation in frequency and severity of cardiovascular and skeletal defects in Sprague-Dawley rats after maternal administration of dimethadione, the N-demethylated metabolite of trimethadione

BACKGROUND: The anticonvulsant trimethadione is a potent inducer of ventricular septation defects, both clinically and in rodents. Teratogenicity requires its N‐demethylation to dimethadione, the proximate teratogen. It was previously demonstrated trimethadione only induced membranous ventricular septation defects in rat (Fleeman et al., 2004 ), and our present goal is to determine whether direct administration of dimethadione increases the incidence and severity of septation defects. METHODS: Pregnant Sprague‐Dawley rats were divided into five groups and administered either distilled water (control) or four different regimens of dimethadione. The core treatment was 300 mg/kg dimethadione b.i.d. on gestation day 9, 10 with additional groups given one additional dose of dimethadione 12 hr earlier, 12 hr later or two additional doses 12 hr earlier and later. Caesarian sections occurred on gestation day 21 and fetuses were examined for standard developmental toxicity endpoints. RESULTS: The broadest dosing regimen yielded the highest incidence and the most severe heart and axioskeletal findings with a decrease in mean fetal body weight. The overall incidence of ventricular septation defects was 74%, of which 68% were membranous and 9% muscular. Outflow tract anomalies (17%) were also observed, as were malformations of the axioskeleton (97%), but not of the long bones, and of particular interest was the high incidence of sternoschesis. CONCLUSIONS: Unlike trimethadione, dimethadione induces more serious muscular septation defects that are believed to be more clinically relevant. This, when taken together with the high incidence of total septation anomalies suggests dimethadione is useful for the study of chemically induced ventricular septation defects. Birth Defects Res (Part B) 92:206–215, 2011. © 2011 Wiley‐Liss, Inc.     

Elucidating protein secondary structures using alpha-carbon recurrence quantifications.

Secondary structures of proteins were studied by recurrence quantification analysis (RQA). High-resolution, 3-dimensional coordinates of alpha-carbon atoms comprising a set of 68 proteins were downloaded from the Protein Data Bank. By fine-tuning four recurrence parameters (radius, line, residue, separation), it was possible to establish excellent agreement between percent contribution of alpha-helix and beta-sheet structures determined independently by RQA and that of the DSSP algorithm (Define Secondary Structure of Proteins). These results indicate that there is an equivalency between these two techniques, which are based upon totally different pattern recognition strategies. RQA enhances qualitative contact maps by quantifying the arrangements of recurrent points of alpha carbons close in 3-dimensional space. For example, the radius was systematically increased, moving the analysis beyond local alpha-carbon neighborhoods in order to capture super-secondary and tertiary structures. However, differences between proteins could only be detected within distances up to about 6-11 A, but not higher. This result underscores the complexity of alpha-carbon spacing when super-secondary structures appear at larger distances. Finally, RQA-defined secondary structures were found to be robust against random displacement of alpha carbons upwards of 1 A. This finding has potential import for the dynamic functions of proteins in motion.