A protein profiling strategy for periodontal disease applications: the Perio-SalivaPRINT

Objectives: It is known that several clinical situations have characteristic molecular deregulations. Some molecular data underlying these deregulations can be found in saliva and have been annotated in databases (SalivaTecDB). Strategies are needed to identify the phenotypes characteristic of these deregulations. Our group has developed a strategy that allows the establishment of saliva protein profiles reflecting different conditions (health and disease). These profiles can be integrated to clinical data (SalivaPRINT Toolkit). The present work aims to identify the Periodontal Diseases (PD)-specific protein profiles. Methods: Unstimulated whole saliva was collected from a group of healthy subjects and a group of PD patients (with gingivitis, periodontitis or periimplantitis). Salivary proteins were separated by the Experion™ automated capillary electrophoresis. The protein profiles of each condition were integrated with the corresponding protein data retrieved from our in-house database (SalivaTecDB). Results: The strategy used enabled the determination of a total protein profile from saliva characteristic of each PDs -the Perio-SalivaPrint. The use of the SalivaPrint Toolkit allowed the identification of molecular weight ranges altered in PD. Using SalivaTecDB we were able to suggest proteins potentially involved in the underlying dysregulated mechanisms of the disease. Conclusions: This approach enabled the determination of a Perio-SalivaPrint – protein profiles specific for gingivitis, periodontitis or periimplantitis - that could empower the use of saliva as a simple and less expensive diagnostic and monitoring fluid. The strategy presented could be an important tool for future applications in the early diagnostic/ screening of Periodontal Disease patients with applications in chairside monitoring.info:eu-repo/semantics/publishedVersio

Prospects and Challenges of Induced Pluripotent Stem Cells in Equine Health

Pluripotent stem cells (PSCs) hold, through the capacity to differentiate into virtually all body cell types, unprecedented promise for human and animal medicine. PSCs are naturally found in the early embryo, and in rodents and humans they can be robustly harvested and grown in culture in the form of embryonic stem cells (ESCs), however the availability of ESCs from horses is limited. ES-like cells named induced pluripotent stem cells (iPSCs) can be derived in vitro by transcription factor-mediated reprogramming of adult cells. As such, iPSCs can be generated in a patient-specific manner providing unmatched potential for tissue transplantation and in vitro disease modelling. In humans, clinical trials using iPSC-derived cells are already taking place and the use of in vitro iPSC models has identified novel mechanisms of disease and therapeutic targets. Although to a more limited extent, iPSCs have also been generated from horses, a species in which, after humans, these cells are likely to hold the greatest potential in regenerative medicine. Before a clinical use can be envisioned, however, significant challenges will need to be addressed in relation to the robust derivation, long-term culture, differentiation and clinical safety of equine iPSCs. Towards this objective, recent studies have reported significant improvement in culture conditions and the successful derivation for the first time of functional cell types from equine iPSCs. Given the wide range of exciting applications they could have, it is hoped future research will make the biomedical promise of iPSCs a reality not only for humans but also horses

Effect of gelation temperature on the properties of skim milk gels made from plant coagulants and chymosin

Reconstituted skim milk was gelled at 25-40°C with the plant-origin coagulants from Cynara cardunculus L. or Cynara humilis L. or with fermentation-produced chymosin. Gel formation and ageing were monitored by low amplitude oscillatory rheology and confocal scanning laser microscopy. Arrhenius plots for the rate of milk gelation were also determined. Plant coagulants had shorter gelation time (tg) at 25°C, 35°C and 40°C, and higher initial rate of increase in G' values at all temperatures tested. The firmest gels at long ageing times were produced by chymosin at 30°C and 32°C. At a gelation temperature of 25°C, the differences in rheological and microstructural characteristics between plant coagulants and chymosin were considerable; plant coagulants had shorter tg and higher G' values. For the lowest gelation temperatures, plant coagulants had smaller activation energy values for gelation. Most of the gelation results were similar between plant coagulants, but some differences were found in the values of tg, the rate of increase in G' and loss tangent parameter. The characteristics of gels produced with plant coagulants were influenced less by the changes in temperature compared with chymosin-produced gels, which may be an important consideration in using plant-origin coagulants in the production of cheeses with a wider range of gelation temperatures.http://www.sciencedirect.com/science/article/B6T7C-493HNG1-1/1/35f20b14e49b2922b16639bac3576d1

P8 - Marine fungi exhibit antimicrobial activity against human oral pathogens

The emergence of resistance to antibiotics and antimycotics has become a challenge in the treatment of infectious diseases, including infections of the oral cavity. Marine fungi are a source of novel biologically active compounds, namely in what concerns the development of antimicrobial and anticancer solutions. Our study aimed to test the antimicrobial activity and the cytotoxicity of the extracts of the two recent identified species of marine fungi, Penicillum lusitanum and Aspergillus affinis. Candida spp. and Enterococcus faecalis isolated from oral pathologies were included to evaluate the antimicrobial potential of the marine fungi by the disk diffusion assay. The cytotoxicity of the effective concentrations of the extract was tested using the Vero cell line (ECACC 88020401, African Green Monkey Kidney cells, GMK clone), according to the ISO 10993-5. The extracts of P. lusitanum and A. affinis were active against C. albicans and E. faecalis, respectively. Penicillum lusitanum active extracts are non-cytotoxic, in contrast to A. affinis extracts that showed high cytotoxic effects on Vero cells, for all concentrations tested. The results on the biological characterization of the P. lusitanumextract are promising and support the development of new disinfecting solutions that may be used during root canal therapy cleaning and shaping.info:eu-repo/semantics/publishedVersio

Stable conditional expression and effect of C/EBPβ-LIP in adipocytes using the pSLIK system

Murine 3T3-L1 adipocytes are widely used as a cellular model of obesity. However, whereas transfection of 3T3-L1 preadipocytes is straightforward, ectopic gene expression in mature 3T3-L1 adipocytes has proved challenging. Here, we used the pSLIK vector system to generate stable doxycycline-inducible expression of the liver-enriched inhibitor protein isoform of CCAAT/enhancer binding protein (C/EBP) {beta} (C/EBP{beta}-LIP) in fully differentiated 3T3-L1 adipocytes. Because overexpression of C/EBP{beta}-LIP impairs adipocyte differentiation, the C/EBP{beta}-LIP construct was first integrated in 3T3-L1 preadipocytes but expression was induced only when adipocytes were fully differentiated. Increased C/EBP{beta}-LIP in mature adipocytes down-regulated C/EBP{beta} target genes including 11{beta}-hydroxysteroid dehydrogenase type 1, phosphoenolpyruvate carboxykinase and fatty acid binding protein 4, but had no effect on asparagine synthetase, demonstrating that transcriptional down-regulation by C/EBP{beta}-LIP in 3T3-L1 adipocytes is not a general effect. Importantly, these genes were modulated in a similar manner in adipose tissue of mice with genetically increased C/EBP{beta}-LIP levels. The use of the pSLIK system to conditionally express transgenes in 3T3-L1 cells could be a valuable tool to dissect adipocyte physiology