Nucleus pulposus phenotypic markers to determine stem cell differentiation : fact or fiction?

Progress in mesenchymal stem cell (MSC) based therapies for nucleus pulposus (NP) regeneration are hampered by a lack of understanding and consensus of the normal NP cell phenotype. Despite the recent consensus paper on NP markers, there is still a need to further validate proposed markers. This study aimed to determine whether an NP phenotypic profile could be identified within a large population of mature NP samples. qRT-PCR was conducted to assess mRNA expression of 13 genes within human non-degenerate articular chondrocytes (AC) (n=10) and NP cells extracted from patients across a spectrum of histological degeneration grades (n=71). qRT-PCR results were used to select NP marker candidates for protein expression analysis. Differential expression at mRNA between AC and non-degenerate NP cells was only observed for Paired Box Protein 1 (PAX1) and Forkhead box F1 (FOXF1). In contrast no other previously suggested markers displayed differential expression between non-degenerate NP and AC at mRNA level. PAX1 and FOXF1 protein expression was significantly higher in the NP compared to annulus fibrosus (AF), cartilaginous endplate (CEP) and AC. In contrast Laminin-5 (LAM-332), Keratin-19 (KRT-19) and Hypoxia Inducible Factor 1 alpha (HIF1α) showed no differential expression in NP cells compared with AC cells. A marker which exclusively differentiates NP cells from AF and AC cells remains to be identified, raising the question: is the NP a heterogeneous population of cells? Or does the natural biological variation during IVD development, degeneration state and even the life cycle of cells make finding one definitive marker impossible

Bacterial antimicrobial metal ion resistance

Metals such as mercury, arsenic, copper and silver have been used in various forms as antimicrobials for thousands of years with until recently, little understanding of their mode of action. The discovery of antibiotics and new organic antimicrobial compounds during the twentieth century saw a general decline in the clinical use of antimicrobial metal compounds, with the exception of the rediscovery of the use of silver for burns treatments and niche uses for other metal compounds. Antibiotics and new antimicrobials were regarded as being safer for the patient and more effective than the metal-based compounds they supplanted. Bacterial metal ion resistances were first discovered in the second half of the twentieth century. The detailed mechanisms of resistance have now been characterized in a wide range of bacteria. As the use of antimicrobial metals is limited, it is legitimate to ask: are antimicrobial metal resistances in pathogenic and commensal bacteria important now? This review details the new, rediscovered and 'never went away' uses of antimicrobial metals; examines the prevalence and linkage of antimicrobial metal resistance genes to other antimicrobial resistance genes; and examines the evidence for horizontal transfer of these genes between bacteria. Finally, we discuss the possible implications of the widespread dissemination of these resistances on re-emergent uses of antimicrobial metals and how this could impact upon the antibiotic resistance problem

Suicide prevention in primary care: General practitioners' views on service availability

BackgroundPrimary care may be a key setting for suicide prevention. However, comparatively little is known about the services available in primary care for suicide prevention. The aims of the current study were to describe services available in general practices for the management of suicidal patients and to examine GPs views on these services. We carried out a questionnaire and interview study in the North West of England. We collected data on GPs views of suicide prevention generally as well as local mental health service provision.FindingsDuring the study period (2003-2005) we used the National Confidential Inquiry Suicide database to identify 286 general practitioners (GPs) who had registered patients who had died by suicide. Data were collected from GPs and practice managers in 167 practices. Responses suggested that there was greater availability of services and training for general mental health issues than for suicide prevention specifically. The three key themes which emerged from GP interviews were: barriers accessing primary or secondary mental health services; obstacles faced when referring a patient to mental health services; managing change within mental health care servicesConclusionsHealth professionals have an important role to play in preventing suicide. However, GPs expressed concerns about the quality of primary care mental health service provision and difficulties with access to secondary mental health services. Addressing these issues could facilitate future suicide prevention in primary care

The association between life events And suicide intent in self-poisoners with and without a history of deliberate self-harm: a preliminary study.

The associations between life events in the 12 months preceding an episode of self-poisoning resulting in hospital attendance (the index episode), and the suicide intent of this episode were compared in individuals for whom the index episode was their first, episode and in individuals in whom it was a recurrence of DSH. Results indicated a significant interaction between independent life events, repetition status, and gender in the prediction of suicide intent, the association between life events and intent being moderated by repetition status in women only. The results provide preliminary evidence to suggest the presence of a suicidal process in women, in which the impact of negative life events on suicide intent diminishes across episodes

Mechanisms of Hsp90 regulation

Heat shock protein 90 (Hsp90) is a molecular chaperone that is involved in the activation of disparate client proteins. This implicates Hsp90 in diverse biological processes that require a variety of co-ordinated regulatory mechanisms to control its activity. Perhaps the most important regulator is heat shock factor 1 (HSF1), which is primarily responsible for upregulating Hsp90 by binding heat shock elements (HSEs) within Hsp90 promoters. HSF1 is itself subject to a variety of regulatory processes and can directly respond to stress. HSF1 also interacts with a variety of transcriptional factors that help integrate biological signals, which in turn regulate Hsp90 appropriately. Because of the diverse clientele of Hsp90 a whole variety of co-chaperones also regulate its activity and some are directly responsible for delivery of client protein. Consequently, co-chaperones themselves, like Hsp90, are also subject to regulatory mechanisms such as post translational modification. This review, looks at the many different levels by which Hsp90 activity is ultimately regulated

High-Throughput Sequencing of RNA Silencing-Associated Small RNAs in Olive (Olea europaea L.)

Small RNAs (sRNAs) of 20 to 25 nucleotides (nt) in length maintain genome integrity and control gene expression in a multitude of developmental and physiological processes. Despite RNA silencing has been primarily studied in model plants, the advent of high-throughput sequencing technologies has enabled profiling of the sRNA component of more than 40 plant species. Here, we used deep sequencing and molecular methods to report the first inventory of sRNAs in olive (Olea europaea L.). sRNA libraries prepared from juvenile and adult shoots revealed that the 24-nt class dominates the sRNA transcriptome and atypically accumulates to levels never seen in other plant species, suggesting an active role of heterochromatin silencing in the maintenance and integrity of its large genome. A total of 18 known miRNA families were identified in the libraries. Also, 5 other sRNAs derived from potential hairpin-like precursors remain as plausible miRNA candidates. RNA blots confirmed miRNA expression and suggested tissue- and/or developmental-specific expression patterns. Target mRNAs of conserved miRNAs were computationally predicted among the olive cDNA collection and experimentally validated through endonucleolytic cleavage assays. Finally, we use expression data to uncover genetic components of the miR156, miR172 and miR390/TAS3-derived trans-acting small interfering RNA (tasiRNA) regulatory nodes, suggesting that these interactive networks controlling developmental transitions are fully operational in olive

Designing microenvironments for optimal outcomes in tissue engineering and regenerative medicine: From biopolymers to culturing conditions

Bone marrow mesenchymal stem cells have been extensively used for tissue engineering and regenerative medicine applications due to their ease of isolation and expansion and their ability to differentiate towards various lineages of mesodermal origin. Despite these properties, their clinical potential is often hampered by the simplicity of the in vitro environment and its inability to resemble the complex in vivo niche. Herein, different microenvironmental cues (e.g. surface topography, substrate stiffness, mechanical stimulation, oxygen tension and co-culture systems) that have been utilised to enhance the therapeutic efficacy of bone marrow mesenchymal stem cells are discussed.The authors would like to acknowledge the following entities for financial support: H2020, Marie Skłodowska-Curie Actions, Innovative Training Networks 2015 Tendon Therapy Train project (Grant No. 676338); Science Foundation Ireland (SFI) / European Regional Development Fund (Grant Number 13/RC/2073); and SFI Career Development Award (Grant Number 15/CDA/3629)

Complex associations between cross‐kingdom microbial endophytes and host genotype in ash dieback disease dynamics

Tree pathogens are a major threat to forest ecosystems. Conservation management strategies can exploit natural mechanisms of resistance, such as tree genotype and host‐associated microbial communities. However, fungal and bacterial communities are rarely looked at in the same framework, particularly in conjunction with host genotype. Here, we explore these relationships and their influence on ash dieback disease, caused by the pathogen Hymenoscyphus fraxineus, in European common ash trees. We collected leaves from UK ash trees and used microsatellite markers to genotype trees, qPCR to quantify H. fraxineus infection load, and ITS and 16S rRNA amplicon sequencing to identify fungal and bacterial communities, respectively. There was a significant association between H. fraxineus infection intensity and ash leaf fungal and bacterial community composition. Higher infection levels were positively correlated with fungal community alpha diversity, and a number of fungal and bacterial genera were significantly associated with infection presence and intensity. Under higher infection loads, leaf microbial networks were characterised by stronger associations between fewer members than those associated with lower infection levels. Together these results suggest that H. fraxineus disrupts stable endophyte communities after a particular infection threshold is reached, and may enable proliferation of opportunistic microbes. We identified three microbial genera associated with an absence of infection, potentially indicating an antagonistic relationship with H. fraxineus that could be utilised in the development of anti‐pathogen treatments. Host genotype did not directly affect infection, but did significantly affect leaf fungal community composition. Thus, host genotype could have the potential to indirectly affect disease susceptibility through genotype x microbiome interactions, and should be considered when selectively breeding trees. Synthesis. We show the diversity, composition and network structure of ash leaf microbial communities are associated with the severity of infection from ash dieback disease, with evidence of disease‐induced dysbiosis. We also show that host genotype influences leaf fungal community composition, but does not directly influence tree infection. These findings help to elucidate relationships between host genetics, the microbiome, and a tree pathogen, highlighting potential resistance mechanisms and possible co‐infection concerns that could inform ash tree manage ment