Proteomic screening of glucose-responsive and glucose non-responsive MIN-6 beta cells reveals differential expression of proteins involved in protein folding, secretion and oxidative stress

The glucose-sensitive insulin-secretion (GSIS) phenotype is relatively unstable in long-term culture of beta cells. The purpose of this study was to investigate relative changes in the proteome between glucose-responsive (low passage) and glucose non-responsive (high passage) murine MIN-6 pancreatic beta cells. The 2D-DIGE and subsequent DeCyder analysis detected 3351 protein spots in the pH range of 4-7. Comparing MIN-6(H) to MIN-6(L) and using a threshold of 1.2-fold, the number of proteins with a decrease in expression level was 152 (4.5%), similar was 3140 (93.7%) and increased 59 (1.8%). From the differentially expressed proteins identified in this study, groups of proteins associated with the endoplasmic reticulum (ER) and proteins involved in oxidative stress were found to be significantly decreased in the high-passage (H passage) cells. These proteins included endoplasmic reticulum protein 29 (ERp29); 78-kDa glucose-related protein, (GRP78); 94-kDa glucose-related protein (GRP94); protein disulphide isomerase; carbonyl reductase 3; peroxidoxin 4 and superoxide dismutase 1. These results suggest that non-GSIS MIN-6 cells do not have the same ability/capacity of glucose-responsive MIN-6 cells to successfully fold, modify or secrete proteins and counteract the problems associated with oxidative stress.     

Periderm- and cortex-based resistance to tuber-feeding Phthorimaea operculella in two wild potato species

The trichome‐bearing wild potatoes Solanum berthaultii (Hawkes) and Solanum tarijense (Hawkes) (Solanaceae) have noted resistance to leaf‐feeding insect herbivores; however, little is known about their resistance to tuber‐feeding herbivores. This study evaluates resistance in tubers of these two species to attack by the potato tuber moth, Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae). Tubers from a range of accessions were presented to recently emerged neonate tuber moth larvae. Resistance to neonates varied between accessions and was generally higher in accessions of S. tarijense. The contribution to observed resistance of periderm vs. cortex factors was assessed by perforating tuber periderm in paired‐tuber experiments. Across species and accessions, an average of 62% of resistance was attributed to periderm‐related factors. All larvae entered tubers through the eyes. Unidentified cortex‐related factors affected larval development time and pupal weight. Sprouting compromised resistance by reducing the protective value of the periderm. The degree of sprouting also decreased larval development times and increased pupal weights in one of two accessions examined. These results demonstrate the potential of S. berthaultii and S. tarijense as sources of tuber‐resistance and identify accessions of both species with notably high periderm‐based protection. Levels of periderm resistance were not correlated to levels of cortex resistance and represent an independent resistance source compatible with the food value of crop potato.     

Evaluation of the site(s) of glycation in human proinsulin by ion-trap LCQ electrospray ionization mass spectrometry

The glycation of beta cell proteins is known to occur under hyperglycemic states. The site(s) of glycation in human proinsulin was investigated following exposure to a hyperglycemic environment under reducing conditions in vitro. Proinsulin and glycated proinsulin were separated by reversed-phase high-performance liquid chromatography (RP-HPLC) and identified using LCQ ion-trap electrospray ionization mass spectrometry. This revealed a major peak (>70% total) of monoglycated proinsulin (M(r) 9552.2 Da), a second peak (approximately 27%) of nonglycated proinsulin (M(r) 9389.8 Da), and a third minor peptide peak (approximately 3%) corresponding to diglycated proinsulin (M(r) 9717.9 Da). Following reduction of disulphide bridges with dithiothreitol, intact peptides were incubated with endoproteinase Glu-C to release nine daughter fragments for LC-MS analysis. This strategy revealed an N-terminal fragment of monoglycated proinsulin Phe(1)-Glu(13), which contained a single glucitol adduct (M(r) 1642.0 Da). A similar treatment of small amounts of purified diglycated proinsulin revealed a fragment with Phe(1)-Glu(13) linked by a disulphide bridge to Gln(70)-Glu(82) containing two glucitol adducts (M(r) 3292.7 Da). In summary, these studies indicate that the major site of glycation in proinsulin, like insulin, is the amino terminal Phe(1) residue. However, small amounts of diglycated proinsulin occur naturally, involving an additional site of glycation located between Gln(70) and Glu(82).     

Axe-Txe,a broad-spectrum proteic toxin-antitoxin system specified by a multidrug-resistant,clinical isolate of Enterococcus faecium

Enterococcal species of bacteria are now acknowledged as leading causes of bacteraemia and other serious nosocomial infections. However, surprisingly little is known about the molecular mechanisms that promote the segregational stability of antibiotic resistance and other plasmids in these bacteria. Plasmid pRUM (24 873 bp) is a multidrug resistance plasmid identified in a clinical isolate of Enterococcus faecium. A novel proteic-based toxin-antitoxin cassette identified on pRUM was demonstrated to be a functional segregational stability module in both its native host and evolutionarily diverse bacterial species. Induced expression of the toxin protein (Txe) of this system resulted in growth inhibition in Escherichia coli. The toxic effect of Txe was alleviated by co-expression of the antitoxin protein, Axe. Homologues of the axe and txe genes are present in the genomes of a diversity of Eubacteria. These homologues (yefM-yoeB) present in the E. coli chromosome function as a toxin-antitoxin mechanism, although the Axe and YefM antitoxin components demonstrate specificity for their cognate toxin proteins in vivo. Axe-Txe is one of the first functional proteic toxin-antitoxin systems to be accurately described for Gram-positive bacteria.     

Endodontic considerations in the elderly

Tooth retention has increased significantly in older adults, and dentists are now challenged by the need to preserve critical teeth. There will be a need to consider endodontic therapy, and this paper describes how successful endodontics can be provided for elderly patients. Strategic treatment planning is essential, and preservation of key teeth will facilitate satisfactory oral function for elderly patients. These teeth may be important in achieving and maintaining an intact anterior dental arch, for removable partial denture retention or preservation of alveolar bone. In some cases, this can only be achieved if endodontic procedures are undertaken. When infection of a root canal is present, there is no reason why good quality endodontic therapy should not work in a healthy elderly patient. Elimination of infection can be challenging in narrow root canals, and a systematic approach for improving access into and negotiating these canals is outlined.     

A statistical measure of tissue heterogeneity with application to 3D PET sarcoma data

In vivo measurement of local tissue characteristics by modern bioimaging techniques such as positron emission tomography (PET) provides the opportunity to analyze quantitatively the role that tissue heterogeneity may play in understanding biological function. This paper develops a statistical measure of the heterogeneity of a tissue characteristic that is based on the deviation of the distribution of the tissue characteristic from a unimodal elliptically contoured spatial pattern. An efficient algorithm is developed for computation of the measure based on volumetric region of interest data. The technique is illustrated by application to data from PET imaging studies of fluorodeoxyglucose utilization in human sarcomas. A set of 74 sarcoma patients (with five-year follow-up survival information) were evaluated for heterogeneity as well as a number of other potential prognostic indicators of survival. A Cox proportional hazards analysis of these data shows that the degree of heterogeneity of the sarcoma is the major risk factor associated with patient death. Some theory is developed to analyze the asymptotic statistical behavior of the heterogeneity estimator. In the context of data arising from Poisson deconvolution (PET being the prime example), the heterogeneity estimator, which is a non-linear functional of the PET image data, is consistent and converges at a rate that is parametric in the injected dose.     

In Vitro and In Vivo Effects of Palmaria palmata Derived Peptides on Glucose Metabolism

International Journal of Peptide Research and Therapeutics - Three synthetic peptides, ILAP, LLAP and MAGVDHI, derived from a Palmaria palmata protein hydrolysate were assessed for their...