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OBJECTIVE: To examine associations and trends between chronic diseases and tooth loss using BRFSS 2012-2018. METHODS: Self-reported permanent tooth loss from tooth decay/gum disease and several self-reported chronic disease diagnoses were analyzed by cycle (2012, 2014, 2016, 2018) to explore associations and trends. Chi-square analyses were performed for the primary outcome of one or more teeth lost with the following ailments: physical health, mental health, weight, diabetes, myocardial infarction, coronary heart disease, stroke, asthma, cancer, respiratory diseases, arthritis, and kidney disease. Multivariate logistic regressions were performed to estimate the odds for tooth-loss for each disease using gender, age, race, insurance, income, education and smoking as covariates. Effects of one or more concurrent chronic disease diagnoses on tooth loss were calculated and 2012-2018 results compared. Interaction between disease and year were used in the multivariate regression aanalyses to find differences in tooth loss from 2012- 2018. All calculations were performed using SAS 9.4. RESULTS: Tooth loss has declined from 45% - in 2012 to 39% - in 2018 in individuals with one chronic disease. A similar decline in tooth loss is seen in those with two, three, four or more chronic diseases. Increased tooth-loss was significantly associated with each chronic disease, with adjusted odds of tooth-loss ranging from 1.08-1.72. Diabetics, had an increased and significant odds of tooth loss with time: 1.36 (2012)-1.54 (2018). The odds of tooth-loss increased as number of concurrent chronic diseases increased -1.2 (one chronic disease)-2.4 (four or more chronic diseases). CONCLUSION: Fewer people are losing teeth, but those with chronic disease experience higher odds of tooth-loss. Having more concurrent diseases is associated with increased tooth-loss. Oral health is essential for overall health, therefore access to oral health care and educating the public and health professionals about these associations is vital.

Chemical modification is a powerful technique for probing functionally important amino acids. N-ethylmaleimide (NEM) reacts readily with exposed sulfhydryl groups, and has previously been shown to inhibit the activity of MT-motor proteins and tubulin assembly. This project seeks to investigate the mechanisms by which NEM affects motor function and inhibits MT minus end assembly. Recombinant motor domains of Drosophila kinesin (DK350 and DK375), Ncd (MC1), and squid kinesin (p181) were modified by NEM. NEM treatment was shown to affect the binding of MC1, but not recombinant kinesin proteins to MTs in the co-sedimentation assay. NEM treatment decreased the MT-stimulated ATPase rates of MC1 and DK350 in an NEM-concentration dependent manner, but did not affect the rate of DK375. Observed effects with DK375, p181, and MC1 were correlated with the number of labeled cysteines determined with [3H]NEM. As previously known, when NEM-treated tubulin was combined with untreated tubulin at certain ratios, assembly occurred only at the MT plus end. To investigate the mechanism by which NEM affects the polarity of tubulin assembly, tubulin was treated with NEM and assembly was analyzed using video-enhanced differential interference contrast microscopy. [3H]NEM was used to follow the time course of modification and to determine the number of modified sites per tubulin subunit. After 10 minutes, one cysteine was labeled on both a and b tubulin and this was sufficient to inhibit minus end assembly. Additionally, having one subunit labeled out of five tubulin subunits was sufficient to observe this effect. Protein digestion methods were used to aid in elimination of cysteines, to characterize potential critical cysteines in MC1, a, and b tubulin.

Ncd is a member of the kinesin family of motor proteins. Ncd is involved in the processes of meiosis and early mitosis in D. melanogaster. PCR-mediated site-directed mutagenesis was utilized to introduce specific mutations into pET/MC6, a construct containing the motor domain of ncd. Six mutations were generated, two at glutamic acid residue 656, two at proline residue 649, one at arginine residue 623, and one double mutant at arginine residue 623 and threonine residue 632. Mutants proteins were expressed in bacteria and further characterized. Mutagenesis of the proline or glutamic acid residues resulted in insoluble proteins. The one exception is the mutagenesis of glutamic acid residue 656 into a glutamine, which resulted in a partially soluble protein. Mutagenesis of the arginine residue into an alanine (MC6-A623) resulted in a soluble protein while the double mutation of the arginine and threonine was insoluble. MC6-A623 exhibited a similar S-sepharose ion exchange chromatography binding and elution profile as MC6. Peptide antibodies made to conserved ncd motor domain sequences also recognized MC6- A623. The affinity of MC6-A623 (under the conditions tested) for microtubules was less than MC6. Most interestingly, under the conditions tested, MC6-A623 did not exhibit an increased ATPase rate in the presence of microtubules, a hallmark of the kinesin family of microtubule motor proteins. Analysis of the published ncd crystal structure, other motor protein sequences, and the experimental results of the mutagenesis of arginine residue 623, suggest that this residue is involved in the binding of MC6 to microtubules.

Drosophila nonclaret disjunctional (Ncd) is a kinesin-like C-terminal motor protein that is involved in spindle assembly in oocytes during meiosis and in spindle maintenance in early embryos during mitosis. Ncd interacts with both "highway" and "cargo" microtubules (MTs) in meiotic and mitotic spindles through the action of ATP-dependent and ATP-independent MT binding sites in the head and tail domains, respectively. Through the action of these binding sites, Ncd bundles and, perhaps, slides MTs relative to each other. These functions are important for the in vivo role of Ncd in the formation of the bipolar spindle and maintenance of the spindle assembly. Despite the high homology of the Ncd head domain to the kinesin head domain, the Ncd tail domain is unique among kinesin-like motor proteins. Characterization of ATP-independent interactions of Ncd with cargo MTs and identification of MT binding sites (located in amino acid residues 83-100 and 115-187) in the tail region by MT co-sedimentation assays revealed that the Ncd tail has functional similarities to microtubule-associated proteins, especially to tau and MAP2, that regulate MT assembly. Like tau MT binding motifs, MT binding sites of the tail domain are rich in basic amino acids that are flanked by proline residues. Cross-linking and MT co-sedimentation assays with subtilisin-digested MTs demonstrated that Ncd tail binding sites (located at the extreme C-terminus and in the H11-H12 loop / H12 helix of each tubulin monomer) on tubulin correspond to tau binding sites. Further, the Ncd tail domain, like tau, can promote and stabilize MT assembly under conditions that induce MT disassembly. Taken together, these results suggest that the Ncd tail functions both in the transport of cargo MTs to spindle poles for the formation of the spindle assembly during meiosis, and in maintenance of spindle assembly during mitosis. How these different functions of Ncd are regulated still remains unknown, however further understanding of the regulation of Ncd function should contribute to our knowledge of cell cycle regulation in both meiotic and mitotic cells.