Development of an in vitro periodontal biofilm model for assessing antimicrobial and host modulatory effects of bioactive molecules

Background: Inflammation within the oral cavity occurs due to dysregulation between microbial biofilms and the host response. Understanding how different oral hygiene products influence inflammatory properties is important for the development of new products. Therefore, creation of a robust host-pathogen biofilm platform capable of evaluating novel oral healthcare compounds is an attractive option. We therefore devised a multi-species biofilm co-culture model to evaluate the naturally derived polyphenol resveratrol (RSV) and gold standard chlorhexidine (CHX) with respect to anti-biofilm and anti-inflammatory properties.<p></p> Methods: An in vitro multi-species biofilm containing <i>S. mitis, F. nucleatum, P. Gingivalis</i> and <i>A. Actinomycetemcomitans</i> was created to represent a disease-associated biofilm and the oral epithelial cell in OKF6-TERT2. Cytotoxicity studies were performed using RSV and CHX. Multi-species biofilms were either treated with either molecule, or alternatively epithelial cells were treated with these prior to biofilm co-culture. Biofilm composition was evaluated and inflammatory responses quantified at a transcriptional and protein level.<p></p> Results: CHX was toxic to epithelial cells and multi-species biofilms at concentrations ranging from 0.01-0.2%. RSV did not effect multi-species biofilm composition, but was toxic to epithelial cells at concentrations greater than 0.01%. In co-culture, CHX-treated biofilms resulted in down regulation of the inflammatory chemokine IL-8 at both mRNA and protein level. RSV-treated epithelial cells in co-culture were down-regulated in the release of IL-8 protein, but not mRNA.<p></p> Conclusions: CHX possesses potent bactericidal properties, which may impact downstream inflammatory mediators. RSV does not appear to have bactericidal properties against multi-species biofilms, however it did appear to supress epithelial cells from releasing inflammatory mediators. This study demonstrates the potential to understand the mechanisms by which different oral hygiene products may influence gingival inflammation, thereby validating the use of a biofilm co-culture model.<p></p&gt

Photoperiod Influences Growth and mll (Mixed-Lineage Leukaemia) Expression in Atlantic Cod

Photoperiod is associated to phenotypic plasticity of somatic growth in several teleost species. However, the molecular mechanisms underlying this phenomenon are currently unknown but it is likely that epigenetic regulation by methyltransferases is involved. The MLL (mixed-lineage leukaemia) family comprises histone methyltransferases that play a critical role in regulating gene expression during early development in mammals. So far, these genes have received scant attention in teleost fish. In the present study, the mean weight of Atlantic cod juveniles reared under continuous illumination was found to be 13% greater than those kept under natural photoperiod conditions for 120 days. We newly determined cDNA sequences of five mll (mll1, mll2, mll3a, mll4b and mll5) and two setd1 (setd1a and setd1ba) paralogues from Atlantic cod. Phylogenetic analysis revealed that the cod genes clustered within the appropriate mll clade and comparative mapping of mll paralogues showed that these genes lie within a region of conserved synteny among teleosts. All mll and setd1 genes were highly expressed in gonads and fast muscle of adult cod, albeit at different levels, and they were differentially regulated with photoperiod in muscle of juvenile fish. Following only one day of exposure to constant light, mll1, mll4b and setd1a were up to 57% lower in these fish compared to the natural photoperiod group. In addition, mRNA expression of myogenic regulatory factors (myog and myf-5) and pax7 in fast muscle was also affected by different photoperiod conditions. Notably, myog was significantly elevated in the continuous illumination group throughout the time course of the experiment. The absence of a day/night cycle is associated with a generalised decrease in mll expression concomitant with an increase in myog transcript levels in fast muscle of Atlantic cod, which may be involved in the observed epigenetic regulation of growth by photoperiod in this species

Gremlin-1 Induces BMP-Independent Tumor Cell Proliferation, Migration, and Invasion

Gremlin-1, a bone morphogenetic protein (BMP) antagonist, is overexpressed in various cancerous tissues but its role in carcinogenesis has not been established. Here, we report that gremlin-1 binds various cancer cell lines and this interaction is inhibited by our newly developed gremlin-1 antibody, GRE1. Gremlin-1 binding to cancer cells was unaffected by the presence of BMP-2, BMP-4, and BMP-7. In addition, the binding was independent of vascular endothelial growth factor receptor-2 (VEGFR2) expression on the cell surface. Addition of gremlin-1 to A549 cells induced a fibroblast-like morphology and decreased E-cadherin expression. In a scratch wound healing assay, A549 cells incubated with gremlin-1 or transfected with gremlin-1 showed increased migration, which was inhibited in the presence of the GRE1 antibody. Gremlin-1 transfected A549 cells also exhibited increased invasiveness as well as an increased growth rate. These effects were also inhibited by the addition of the GRE1 antibody. In conclusion, this study demonstrates that gremlin-1 directly interacts with cancer cells in a BMP- and VEGFR2-independent manner and can induce cell migration, invasion, and proliferation

Variation in Phenotype, Parasite Load and Male Competitive Ability across a Cryptic Hybrid Zone

BackgroundMolecular genetic studies are revealing an increasing number of cryptic lineages or species, which are highly genetically divergent but apparently cannot be distinguished morphologically. This observation gives rise to three important questions: 1) have these cryptic lineages diverged in phenotypic traits that may not be obvious to humans; 2) when cryptic lineages come into secondary contact, what are the evolutionary consequences: stable co-existence, replacement, admixture or differentiation and 3) what processes influence the evolutionary dynamics of these secondary contact zones?Methodology/principal findingsTo address these questions, we first tested whether males of the Iberian lizard Lacerta schreiberi from two highly genetically divergent, yet morphologically cryptic lineages on either side of an east-west secondary contact could be differentiated based on detailed analysis of morphology, coloration and parasite load. Next, we tested whether these differences could be driven by pre-copulatory intra-sexual selection (male-male competition). Compared to eastern males, western males had fewer parasites, were in better body condition and were more intensely coloured. Although subtle environmental variation across the hybrid zone could explain the differences in parasite load and body condition, these were uncorrelated with colour expression, suggesting that the differences in coloration reflect heritable divergence. The lineages did not differ in their aggressive behaviour or competitive ability. However, body size, which predicted male aggressiveness, was positively correlated with the colour traits that differed between genetic backgrounds.Conclusions/significanceOur study confirms that these cryptic lineages differ in several aspects that are likely to influence fitness. Although there were no clear differences in male competitive ability, our results suggest a potential indirect role for intra-sexual selection. Specifically, if lizards use the colour traits that differ between genetic backgrounds to assess the size of potential rivals or mates, the resulting fitness differential favouring western males could result in net male-mediated gene flow from west to east across the current hybrid zone.Devi Stuart-Fox, Raquel Godinho, Joëlle Goüy de Bellocq, Nancy R. Irwin, José Carlos Brito, Adnan Moussalli, Pavel Široký, Andrew F. Hugall and Stuart J. E. Bair

Male Weaponry in a Fighting Cricket

Sexually selected male weaponry is widespread in nature. Despite being model systems for the study of male aggression in Western science and for cricket fights in Chinese culture, field crickets (Orthoptera, Gryllidae, Gryllinae) are not known to possess sexually dimorphic weaponry. In a wild population of the fall field cricket, Gryllus pennsylvanicus, we report sexual dimorphism in head size as well as the size of mouthparts, both of which are used when aggressive contests between males escalate to physical combat. Male G. pennsylvanicus have larger heads, maxillae and mandibles than females when controlling for pronotum length. We conducted two experiments to test the hypothesis that relatively larger weaponry conveys an advantage to males in aggressive contests. Pairs of males were selected for differences in head size and consequently were different in the size of maxillae and mandibles. In the first experiment, males were closely matched for body size (pronotum length), and in the second, they were matched for body mass. Males with proportionately larger weaponry won more fights and increasing differences in weaponry size between males increased the fighting success of the male with the larger weaponry. This was particularly true when contests escalated to grappling, the most intense level of aggression. However, neither contest duration nor intensity was related to weaponry size as predicted by models of contest settlement. These results are the first evidence that the size of the head capsule and mouthparts are under positive selection via male-male competition in field crickets, and validate 800-year-old Chinese traditional knowledge

Divergent receiver responses to components of multimodal signals in two foot-flagging frog species.

Multimodal communication of acoustic and visual signals serves a vital role in the mating system of anuran amphibians. To understand signal evolution and function in multimodal signal design it is critical to test receiver responses to unimodal signal components versus multimodal composite signals. We investigated two anuran species displaying a conspicuous foot-flagging behavior in addition to or in combination with advertisement calls while announcing their signaling sites to conspecifics. To investigate the conspicuousness of the foot-flagging signals, we measured and compared spectral reflectance of foot webbings of Micrixalus saxicola and Staurois parvus using a spectrophotometer. We performed behavioral field experiments using a model frog including an extendable leg combined with acoustic playbacks to test receiver responses to acoustic, visual and combined audio-visual stimuli. Our results indicated that the foot webbings of S. parvus achieved a 13 times higher contrast against their visual background than feet of M. saxicola. The main response to all experimental stimuli in S. parvus was foot flagging, whereas M. saxicola responded primarily with calls but never foot flagged. Together these across-species differences suggest that in S. parvus foot-flagging behavior is applied as a salient and frequently used communicative signal during agonistic behavior, whereas we propose it constitutes an evolutionary nascent state in ritualization of the current fighting behavior in M. saxicola