Search

Uterus tissue engineering may dismantle limitations in current uterus transplantation protocols. A uterine biomaterial populated with patient-derived cells could potentially serve as a graft to circumvent complicated surgery of live donors, immunosuppressive medication and rejection episodes. Repeated uterine bioengineering studies on rodents have shown promising results using decellularised scaffolds to restore fertility in a partially impaired uterus and now mandate experiments on larger and more human-like animal models. The aim of the presented studies was therefore to establish adequate protocols for scaffold generation and prepare for future in vivo sheep uterus bioengineering experiments. Three decellularisation protocols were developed using vascular perfusion through the uterine artery of whole sheep uteri obtained from slaughterhouse material. Decellularisation solutions used were based on 0.5% sodium dodecyl sulphate (Protocol 1) or 2% sodium deoxycholate (Protocol 2) or with a sequential perfusion of 2% sodium deoxycholate and 1% Triton X-100 (Protocol 3). The scaffolds were examined by histology, extracellular matrix quantification, evaluation of mechanical properties and the ability to support foetal sheep stem cells after recellularisation. We showed that a sheep uterus can successfully be decellularised while maintaining a high integrity of the extracellular components. Uteri perfused with sodium deoxycholate (Protocol 2) were the most favourable treatment in our study based on quantifications. However, all scaffolds supported stem cells for 2 weeks in vitro and showed no cytotoxicity signs. Cells continued to express markers for proliferation and maintained their undifferentiated phenotype. Hence, this study reports three valuable decellularisation protocols for future in vivo sheep uterus bioengineering experiments.

The dwarf succulent genus Lithops is endemic to Southern Africa and of considerable conservation concern. Species delimitation is often problematic and based mainly on leaf morphology, which is strongly associated with habitat. Relationships between taxa and populations in Namibia were studied with amplified fragment length polymorphisms using 44 wild Lithops populations representing 15 species and 23 taxa. Four primer pairs produced 92 polymorphic bands in the 223 samples. Expected heterozygosity (He) within taxa ranged from 0.086 to 0.450. Genetic and geographic distances were correlated according to a Mantel test. Analysis of molecular variance showed only 23% variation among the 15 investigated species. Genetic differentiation and structuring were investigated with a principal coordinate analysis, a neighbour-joining and a Bayesian phylogeny, a Bayesian clustering analysis and a discriminant analysis of principal components. In all five analyses, L. optica and L. herrei, which differ only in flower colour, clustered closely together and are here combined under L. optica. The morphologically similar species L. amicorum and L. karasmontana clustered together. Lithops amicorum is therefore reduced to subspecific level: L. karasmontana subsp. amicorum, comb. nov. Subspecific taxa overlapped to a large extent except in L. karasmontana where 13% of the variability resided among subspecies, whereas the nominal subspecies differed from subsp. bella and subsp. eberlanzii; the latter two could not be separated and are here combined under L. karasmontana subsp. bella.

Uterine factor infertility was considered incurable until recently when we reported the first successful live birth after uterus transplantation. However, risky donor surgery and immunosuppressive therapy are factors that may be avoided with bioengineering. For example, transplanted recellularized constructs derived from decellularized tissue restored fertility in rodent models and mandate translational studies. In this study, we decellularized whole sheep uterus with three different protocols using 0.5% sodium dodecyl sulfate, 2% sodium deoxycholate (SDC) or 2% SDC, and 1% Triton X-100. Scaffolds were then assessed for bioactivity using the dorsal root ganglion and chorioallantoic membrane assays, and we found that all the uterus scaffolds exhibited growth factor activity that promoted neurogenesis and angiogenesis. Extensive recellularization optimization was conducted using multipotent sheep fetal stem cells and we report results from the following three in vitro conditions; (a) standard cell culturing conditions, (b) constructs cultured in transwells, and (c) scaffolds preconditioned with matrix metalloproteinase 2 and 9. The recellularization efficiency was improved short-term when transwells were used compared with standard culturing conditions. However, the recellularization efficiency in scaffolds preconditioned with matrix metalloproteinases was 200–300% better than the other strategies evaluated herein, independent of decellularization protocol. Hence, a major recellularization hurdle has been overcome with the improved recellularization strategies and in vitro platforms described herein. These results are an important milestone and should facilitate the production of large bioengineered grafts suitable for future in vivo applications in the sheep, which is an essential step before considering these principles in a clinical setting.

Immunoglobulin G (IgG) is important in clearance and recognition of previously presented antigens and after activation, IgGs can interact with the Fc gamma receptors (FcγRs) on haematopoietic cells, including bone‐resorbing osteoclasts. The pathogenicity of IgG, that is the ability to elicit stimulatory effects via FcγRs, can be modulated by attachment of sugar moieties, including sialic acids. Human IgGs and autoantibodies are associated with bone loss in autoimmune disease. However, the impact of polyclonal murine IgG via FcγRs on bone loss is poorly understood. Here, we investigate if heat‐aggregated activated murine polyclonal IgG complexes have any direct effects on murine osteoclasts and if they modulate arthritis‐mediated bone loss. Using cell cultures of murine osteoclasts, we show that IgG complexes without sialic acids (de‐IgG complexes) enhance receptor activator of nuclear factor kappa‐Β ligand (RANKL)‐stimulated osteoclastogenesis, an effect associated with increased FcγRIII expression. Using an in vivo model of arthritis‐mediated bone loss, where IgG complexes were injected into arthritic knees, no effect on the severity of arthritis or the degree of arthritis‐mediated bone loss was detected. Interestingly, injection of de‐IgG complexes into non‐arthritic knees increased osteoclast formation and enhanced bone erosions. Our findings show that activated de‐IgG complexes have no additive effect on arthritis‐mediated bone loss. However, de‐IgG complexes potentiate murine osteoclastogenesis and enhance local bone erosion in non‐arthritic bones, further confirming the link between the adaptive immune system and bone.

Non-invasive electrical stimulation (ES) is increasingly applied to improve vision in untreatable eye conditions, such as retinitis pigmentosa and age-related macular degeneration. Our previous study suggested that ES promoted retinal function and the proliferation of progenitor-like glial cells in mice with inherited photoreceptor degeneration, however, the underlying mechanism remains obscure. M&uuml, ller cells (MCs) are thought to be dormant residential progenitor cells that possess a high potential for retinal neuron repair and functional plasticity. Here, we showed that ES with a ramp waveform of 20 Hz and 300 &micro, A of current was effective at inducing mouse MC proliferation and enhancing their expression of progenitor cell markers, such as Crx (cone&ndash, rod homeobox) and Wnt7, as well as their production of trophic factors, including ciliary neurotrophic factor. RNA sequencing revealed that calcium signaling pathway activation was a key event, with a false discovery rate of 5.33 &times, 10&minus, 8 (P = 1.78 &times, 10) in ES-mediated gene profiling changes. Moreover, the calcium channel blocker, nifedipine, abolished the observed effects of ES on MC proliferation and progenitor cell gene induction, supporting a central role of ES-induced Ca2+ signaling in the MC changes. Our results suggest that low-current ES may present a convenient tool for manipulating MC behavior toward neuroregeneration and repair.

The genotyping of European fruit tree collections has helped to identify synonyms, determine parentage, reveal key specimens in the collections and provide information on the development of modern cultivars from one or several progenitors. However, studies on European plum Prunus domestica L. accessions have been lagging behind, mainly because of the hexaploid chromosome number. In this co-operative study, 104 accessions conserved by 14 partners across Europe were phenotyped for 20 descriptors, and genotyped for 8 SSR loci together with 8 reference cultivars. Based on the descriptors and additional information supplied by the partners, as well as the scientific and horticultural literature, each accession was assigned to one of six pomological groups; (1) egg plums sensu lato (E), (2) prunes of the French d’Agen type (P/A), (3) prunes of the Central-Southeast European Zwetschen type (P/Z), (4) greengages (G), (5) mirabelles (M) and (6) bullaces, damsons and var. pomariorum (D/B). A MANOVA conducted on descriptor data revealed significant differentiation among the pomological groups as well as a geographic impact on the differentiation of local plum accessions in Europe. SSR data showed that two trios and seven pairs of genotypes had very similar allele profiles and possibly are genetically identical in spite of different accession names. An AMOVA indicated sparse genetic differentiation when accessions were grouped according to geographic origin whereas significant differences were obtained among pomological groups. A Bayesian analysis of genetic structure, as well as a discriminant analysis of principal components (DAPC), further revealed levels of similarity among and within the different pomological groups, suggesting that egg plums sensu lato (E) and greengages (G) can be referred to subsp. domestica while damsons and bullaces (D/B) but also Central-Southeast European prunes (P/Z) show more affinity to subsp. insititia. The small and possibly heterogeneous groups with mirabelles (M) and prunes of the d’Agen type (P/A) take an intermediate position suggesting a hybridogenic origin.

Growing pear has a long tradition in Tunisia, and numerous local cultivars possessing an excellent adaptability and resilience potential to climatic variation are present. This large adaptability is associated with an important genetic diversity, which is threatened to erosion. Appropriate measures have to be taken in order to properly evaluate and conserve this local material. Microsatellite markers were used to assess the level of genetic diversity among Tunisian pear germplasm, and compare it with some European varieties and wild pear species. 61 pear accessions representing eight groups (six groups from Tunisia, one from Northern Europe and another group composed of wild pear) have been genotyped using SSR markers derived from apple and pear. The pear accessions showed a significant polymorphism and 95 polymorphic alleles were found. The number of alleles per locus varied from 5 for CH04e03 locus to 14 for CH01d09 locus with an average of 9.4 alleles per locus. Moreover, the mean gene diversity (He) per locus ranged from 0.192 to 0.752. Genetic distance values and cluster analyses revealed high genetic similarities among the Tunisian groups. Factorial correspondence analysis (FCA) categorized the accessions into three independent groups where Tunisian local accessions agglomerated together distantly from European and wild pear accessions. Additionally, UPGMA dendrogram grouped accessions into two clusters, confirmed thereafter by the Bayesian model-based Structure analysis. The results showed 16 putative triploid accessions found in the local germplasm. This study provides valuable information to develop strategies of local pear conservation and use. Keywords: Pyrus communis, molecular marker, genetic diversity, local accession

MicroRNAs (miRNAs) are non-coding RNAs that are involved in various biological pathways by regulating gene expression. Teeth develop via reciprocal and sequential interactions between the epithelium and the ectomesenchyme. The speci.c functions of several genes during tooth development are known, and the involvement of their mutations in the pathogenesis of congenital dental defects has been widely studied. The miRNA pathway is considered to have a significant role in embryogenesis including tooth development. It has been shown that miRNAs regulate morphogenesis of tooth by fine-tuning the signalling networks, however, their precise role in tooth differentiation and morphogenesis is still elusive. The present review focuses on the studies that have used animal models to explore the function of miRNAs in tooth development. Major findings with special emphasis on the miRNA involvement in .ne-tuning and network regulation are presented and discussed. Disturbances in tooth development in the global miRNA processing knockouts mirror the essential fine-tuning guiding appropriate formation of dental hard tissues. However, further investigation of single miRNA function and mutation, including deletion and overexpression, may lead to improved knowledge on development of particular dental defects in humans. In the light of similarities between tooth development and other organs originating from the epithelium, further understanding of miRNAs` function during tooth development may have wide biological relevance.

Background: The miR-106a-363 cluster, encoding six miRNAs (miR- 106a, miR-18b, miR-20b, miR-19b-2, miR-92-2 and miR-363), has been shown to be overexpressed in various tumours. In oral carcinoma cells, however only miR- 106a was detectable from this cluster. We have investigated how effects of transfection of oral carcinoma cells with a non-expressed member of this cluster affect mRNA transcriptomes and cellular selected functions. Methods: Investigate effects of miR-20b and miR-363 mimics on cellular respiration, glycolysis and mobility. Effects on mRNA transcriptomes were monitored using microarrays. Results: The studies show that in oral carcinoma cells transfected with miR-20b -, or miR-363-3p or miR-363-5p mimic different mRNAs were differentially expressed. Nevertheless, bioinformatics analysis suggested significant associations of differentially expressed genes to inhibition of cellular proliferation, cell cycle and cellular migration. These results were also experimentally confirmed. Conclusions: Transfection of miRNA mimics for unexpressed members of the miR-106a-363 cluster (miR20b, miR-363-3p and miR-363-5p) exhibit an anti-proliferative effect on oral carcinoma cells, although likely mediated by different regulatory mechanisms.

Acute bacterial parotitis is uncommon in young adults. Infection with Serratia marcescens is even rarer and usually found in hospitalized patients. This case report focuses on a young woman with acute bacterial parotitis caused by S. marcescens that required a longer-than-normal course of antibiotics.

Cultivated apple is one of the most widely grown fruit crops worldwide. With the introduction of modern apple cultivars, from foreign and national breeding programs, the use of local cultivars decreased during the 20th century. In order to minimize genetic erosion and avoid loss of special genotypes, a number of local clonal archives were established across Sweden, with the goal of retaining old and local cultivars. About 220 apple cultivars, appointed for preservation, obtained the status of mandate cultivars. Initially, they were identified based on pomological traits, but prior to the establishment of the Swedish Central Collection they were genotyped with simple sequence repeat (SSR) markers. SSR markers helped to evaluate the status of the preserved material, as well as to find the best possible true-to-type source for propagation, thus guiding the establishment of the Central Collection. Recently, 215 accessions from this collection were genotyped using the 20 K apple Infinium® single nucleotide polymorphism (SNP) array, in order to gain insight into its genetic structure. The initial SSR analysis confirmed the identity of multiple samples with the same cultivar name grown in different locations and identified several mislabeled samples. In the subsequent SNP analysis we identified 30 clonal relationships and a number of parent-offspring relationships, including 18 trios. We also identified five cultivar samples with inconsistent ploidy levels between the SNP and SSR data, in some cases indicating problematic samples preserved in either the Central Collection or some of the local clonal archives. These cultivars need further investigation to ensure their true-to-typeness. Furthermore, the Swedish Central Collection has continued to grow since the onset of this work and now contains additional cultivars, which should be included in future studies. The results indicate that a number of the preserved mandate cultivars holds high potential value for modern breeding programs.

a b s t r a c t Different approaches for germplasm conservation can have considerable impact on the amount of genetic diversity captured. Nine SSR loci were used to compare genetic diversity and genetic structure of tradi- tional Nordic plum accessions maintained using two different approaches; ex situ (germplasm collection, Balsgard in Sweden), and on-farm (Southern Norway). In addition, eight international reference cultivars were included. A total of 204 distinct SSR alleles were amplified, revealing high overall diversity. Only one duplicate was found in the ex situ collection while numerous duplicates were found in the on-farm material. There was little differentiation between the three groups of samples, i.e. the Swedish ex situ col- lection, the Norwegian on-farm collection and the international reference cultivars. However, a Bayesian analysis of genetic structure revealed a clear difference in genetic structure between the Swedish ex situ collection and the Norwegian on-farm collection. After classifying the ex situ plum accessions into four different morphological groups/species, based on their pedigree data and phenotype, a more pronounced clustering was noted. The results of the Bayesian analyses thus indicated that 'Krikon' (subsp. insititia) and some possibly related genotypes from Norway form the most divergent cluster. Other morphological groups/species that could be discerned although with some overlap, are greengages (e.g. 'Reine Claude'), common European plum (exemplified with 'Victoria' and 'Stanley') and two Norwegian accessions with unknown affinity. © 2015 Elsevier B.V. All rights reserved.

The circadian clock is comprised of a master component situated in the hypothalamic suprachiasmatic nucleus and subordinate clock genes in almost every cell of the body. The circadian clock genes and their encoded proteins govern the organism to follow the natural signals of time, and adapt to external changes in the environment. The majority of physiological processes in mammals exhibit variable circadian rhythms, which are generated and coordinated by an oscillation in the expression of the clock genes. A number of studies have reported that alteration in the expression level of clock genes is correlated with several pathological conditions, including cancer. However, little is known about the role of clock genes in homeostasis of the oral epithelium and their disturbances in oral carcinogenesis. The present review summarizes the current state of knowledge of the implications of clock genes in oral cancer. It has been demonstrated that the development of oral squamous cell carcinoma undergoes circadian oscillation in relation to tumor volume and proliferation rate. The circadian clock gene period (PER)1 has been associated with oral cancer pathogenesis and it is suggested that changes in the expression of PER1 may exhibit an important role in the development, invasion, and metastasis of oral squamous cell carcinoma. However, its role remains elusive and there is a need for further research in order to understand the underlying mechanisms of the clock genes in oral cancer pathogenesis.

The genus Harpagophytumhas two species: H. procumben swhich is an important medicinal plant in southern Africa, and H. zeyheri. Genetic diversity in 96 samples, obtained by germinating seeds collected from Botswana, was assessed using six inter-simple sequence repeat (ISSR) and 10 random amplified polymorphic DNA (RAPD) primers. These DNA markers yielded a total of 138 polymorphic bands. Polymorphism information content (PIC) ranged from 0.06 to 0.39 for ISSR primers, and from 0.09 to 0.43 for RAPD primers. Jaccard's similarity coefficients were highest when seedlings derived from the same fruit capsule were compared, while seedlings from different fruits on the same plant had intermediate values. The lowest values were recorded among seedlings from different plants. These results were consistent with an outcrossing breeding system in Harpagophytum. Analysis of molecular variance revealed significant differentiation (P < 0.01) between taxonomic units within Harpagophytum. About 39% of the variability occurred between the two species, H. procumbens and H. zeyheri. Plants with an intermediate morphology, i.e. putative hybrids (PH), showed 21% differentiation when compared with H. procumbens ssp. procumbens (PP), and 19% when compared with H. procumbens ssp. transvaalense (PT) or with H. zeyheri (ZZ). In addition, a deviating variant of PT was identified, here termed ‘procumbens new variety’ (PN). PN showed only 9% differentiation when compared with PT, 22% when compared with PP or with PH, and 41% when compared with ZZ. Considerable differentiation between the two Harpagophytum species was revealed also by a cluster analysis. Introgression was, however, suggested by the intermediate position of the putative hybrid plants in a principal component analysis while inter-specific gene flow was shown by a Bayesian genetic structure analysis.

Incremental markings in dental enamel suggest that the circadian clock may influence the molecular underpinnings orchestrating enamel formation. The aim of this study was to investigate whether the genes and microRNAs (miRNAs) oscillate in a circadian pattern during tooth and enamel development.Comparative gene and miRNA expression profiling of the first mandibular molar tooth germ isolated at different time-points during the light and night period was performed using microarrays and validated using real-time RT-PCR. Bioinformatic analysis was carried out using Ingenuity Pathway Analysis (IPA), and TargetScan software was used in order to identify computationally predicted miRNA-mRNA target relationships.In total, 439 genes and 32 miRNAs exhibited significantly different (p 0.05) levels of expression in the light phase compared with the night phase tooth germs. Genes involved in enamel formation, i.e. Amelx, Ambn, Amtn, and Odam, oscillated in a circadian pattern. Furthermore, the circadian clock genes, in particular Clock and Bmal1, oscillated in mouse molar tooth germ during 24-h intervals. The expression of Clock and Bmal1 was inversely correlated with the expression of miR-182 and miR-141, respectively.MiRNAs, including miR-182 and miR-141, are involved in the control of peripheral circadian rhythms in the developing tooth by regulating the expression of genes coding for circadian transcription factors such as CLOCK and BMAL1. Regulation of circadian rhythms may be important for enamel phenotype, and the morphology of dental enamel may vary between individuals due to differences in circadian profiles.

This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Purpose Transplantation of limbal stem cells is a promising therapy for limbal stem cell deficiency. Limbal cells can be harvested from either a healthy part of the patient’s eye or the eye of a donor. Small explants are less likely to inflict injury to the donor site. We investigated the effects of limbal explant size on multiple characteristics known to be important for transplant function. Methods Human limbal epithelial cells were expanded from large versus small explants (3 versus 1 mm of the corneal circumference) for 3 weeks and characterized by light microscopy, immunohistochemistry, and transmission electron microscopy. Epithelial thickness, stratification, outgrowth, ultrastructure and phenotype were assessed. Results Epithelial thickness and stratification were similar between the groups. Outgrowth size correlated positively with explant size (r = 0.37; P = 0.01), whereas fold growth correlated negatively with explant size (r = –0.55; P < 0.0001). Percentage of cells expressing the limbal epithelial cell marker K19 was higher in cells derived from large explants (99.1±1.2%) compared to cells derived from small explants (93.2±13.6%, P = 0.024). The percentage of cells expressing ABCG2, integrin β1, p63, and p63α that are markers suggestive of an immature phenotype; Keratin 3, Connexin 43, and E-Cadherin that are markers of differentiation; and Ki67 and PCNA that indicate cell proliferation were equal in both groups. Desmosome and hemidesmosome densities were equal between the groups. Conclusion For donor- and culture conditions used in the present study, large explants are preferable to small in terms of outgrowth area. As regards limbal epithelial cell thickness, stratification, mechanical strength, and the attainment of a predominantly immature phenotype, both large and small explants are sufficient.

The developing tooth can be regarded as a typical vertebrate organ starting as an epithelial bud that subsequently develops into a mature organ, a process that entails extensive epithelial-mesenchymal interactions. Functions of various signaling molecules and gene expression during odontogenesis have appealed considerable interest. Although it now is evident that microRNAs (miRNAs) are implicated in tooth development, their regulative role remains essentially unknown. MicroRNAs have been studied in the context of most ectodermal organs but only a few functional studies of their role in tooth development have been published so far. The fine-tuning of this epithelial-mesenchymal network via the

A set of 85 heirloom apple cultivars aimed for long-term preservation in two germplasm collections in Sweden and Finland was evaluated with 8 SSR primer pairs to evaluate genetic diversity and genetic relatedness. An additional set of 16 European cultivars was included for comparison. The eight SSR primer pairs amplified 9 loci and 105 alleles. Genetic analyses performed by MDS indicated some differentiation between Swedish and Finnish cultivars, with European cultivars intermixed with the Swedish. The existence of three groups was, however, indicated by a Bayesian model-based clustering. One of the groups was clearly dominated by Swedish cultivars and another by Finnish. The third group included almost equal proportions of representatives from all three areas. The obtained results confirmed the genetic distinctness of Finnish apple cultivars, which can be explained by climate adaptation and admixture with a Russian gene pool.

Fifteen expressed sequence tag (EST)-derived simple sequence repeats (EST-SSRs) were used to investigate genetic diversity in 139 plants obtained from seeds of 35 watermelon accessions collected from all the geographical provinces of Zimbabwe. In addition, 15 plants representing three commercial varieties developed in the United States (USA) were analyzed for comparison. A total of 65 alleles were detected among all the watermelon accessions. For the 13 polymorphic EST-SSR loci, number of alleles per locus varied from 2 to 13, with an average of 5 alleles per locus. Values for the polymorphic information content increased as the number of alleles increased, and varied from 0.15 to 0.77 with an average of 0.54 suggesting sufficient discriminatory power. Both cluster analysis and principal coordinate analysis (PCA) produced two major clusters; one with the 22 cow-melon accessions and the other with the 16 sweet watermelon accessions. Within the sweet watermelon group, two distinct sub-clusters formed, one of which contained only two of the commercial varieties from USA. Partitioning of genetic variation in the Zimbabwean material using analysis of molecular variation (AMOVA) revealed that 64% of the total variation resides between the two major forms, i.e. sweet watermelons and cow-melons, 28% between accessions within forms and 8% within accessions. The EST-SSR markers revealed a somewhat higher diversity in sweet watermelon accessions compared to that of cow-melons. This finding is contrary to previous reports using other markers (genomic SSR loci or RAPD) and/or a plant material that is likely to have experienced more stringent selection procedures compared to the landraces analyzed in our study.

MicroRNAs (miRNAs) are a class of small, non-coding RNAs that provide an efficient pathway for regulation of gene expression at a post-transcriptional level. Tooth development is regulated by a complex network of cell-cell signaling during all steps of organogenesis. Most of the congenital dental defects in humans are caused by mutations in genes involved in developmental regulatory networks. Whereas the developmental morphological stages of the tooth development already are thoroughly documented, the implicated genetic network is still under investigation. The involvement of miRNAs in the regulation of tooth genetic network was suggested for the first time in 2008. MiRNAs regulate tooth morphogenesis by fine-tuning the signaling networks. Unique groups of miRNAs are expressed in dental epithelium compared with mesenchyme, as well as in molars compared with incisors. The present review focuses on the current state of knowledge on the expression and function of miRNAs in human dental tissues, including teeth and the surrounding structures. Herein, we show that miRNAs exhibit specific roles in human dental tissues and are involved in gingival and periodontal disease, tooth movement and eruption, dental pulp physiology including repair and regeneration, differentiation of dental cells, and enamel mineralization. In light of similarities between the tooth development and other organs originating from the epithelium, further understanding of miRNAs` function in dental tissues may have wide biological relevance.

Purpose Storage of cultured human oral keratinocytes (HOK) allows for transportation of cultured transplants to eye clinics worldwide. In a previous study, one-week storage of cultured HOK was found to be superior with regard to viability and morphology at 12 degrees C compared to 4 degrees C and 37 degrees C. To understand more of how storage temperature affects cell phenotype, gene expression of HOK before and after storage at 4 degrees C, 12 degrees C, and 37 degrees C was assessed. Materials and Methods Cultured HOK were stored in HEPES-and sodium bicarbonate-buffered Minimum Essential Medium at 4 degrees C, 12 degrees C, and 37 degrees C for one week. Total RNA was isolated and the gene expression profile was determined using DNA microarrays and analyzed with Partek Genomics Suite software and Ingenuity Pathway Analysis. Differentially expressed genes (fold change > 1.5 and P < 0.05) were identified by one-way ANOVA. Key genes were validated using qPCR. Results Gene expression of cultures stored at 4 degrees C and 12 degrees C clustered close to the unstored control cultures. Cultures stored at 37 degrees C displayed substantial change in gene expression compared to the other groups. In comparison with 12 degrees C, 2,981 genes were differentially expressed at 37 degrees C. In contrast, only 67 genes were differentially expressed between the unstored control and the cells stored at 12 degrees C. The 12 degrees C and 37 degrees C culture groups differed most significantly with regard to the expression of differentiation markers. The Hedgehog signaling pathway was significantly downregulated at 37 degrees C compared to 12 degrees C. Conclusion HOK cultures stored at 37 degrees C showed considerably larger changes in gene expression compared to unstored cells than cultured HOK stored at 4 degrees C and 12 degrees C. The changes observed at 37 degrees C consisted of differentiation of the cells towards a squamous epithelium-specific phenotype. Storing cultured ocular surface transplants at 37 degrees C is therefore not recommended. This is particularly interesting as 37 degrees C is the standard incubation temperature used for cell culture. Funding Agencies|Department of Oral Biology, Faculty of Dentistry, University of Oslo; Department of Medical Biochemistry, Oslo University Hospital, Oslo, Norway

MicroRNAs (miRNAs) form a class of noncoding RNA genes whose products are small single-stranded RNAs that are involved in the regulation of translation and degradation of mRNAs. There is a fine balance between deregulation of normal developmental programs and tumor genesis. An increasing body of evidence suggests that altered expression of miRNAs is entailed in the pathogenesis of human cancers. Studies in mouse and human cells have identified the miR-17-92 cluster as a potential oncogene. The miR-17-92 cluster is often amplified or overexpressed in human cancers and has recently emerged as the prototypical oncogenic polycistron miRNA. The functional analysis of miR-17-92 is intricate by the existence of two paralogues: miR-106a-363 and miR-106b-25. During early evolution of vertebrates, it is likely that the three clusters commenced via a series of duplication and deletion occurrences. As miR-106a-363 and miR-106b-25 contain miRNAs that are very similar, and in some cases identical, to those encoded by miR-17-92, it is feasible that they regulate a similar set of genes and have overlapping functions. Further understanding of these three clusters and their functions will increase our knowledge about cancer progression. The present review discusses the characteristics and functions of these three miRNA clusters.

Simple sequence repeat (SSR) markers are increasingly being used to confirm the identification of accessions in plant collections and/or to quantify their relatedness. However, the possibility to compare SSR data in different databases, is dependent on the use of identical SSR loci. Therefore, a standard set of 17 SSR loci was recently appointed by the European Cooperative Program for Plant Genetic Resources (ECPGR) for the screening of pear accessions. Here, we report the results from using 10 of these loci to fingerprint a total of 86 samples from Swedish pear collections including 49 mandate (heritage) cultivars, and 8 reference genotypes from Brogdale. The ability to compare our data with previously obtained data from fingerprinting the Brogdale collection with the same loci, allowed us to verify the labelling of some cultivars as well as to detect several synonyms and erroneous labellings. Obvious cases of mislabelling affected 14 of the analysed cultivars, i.e., 29%. A principal coordinates (PCO) analysis indicated that a loosely defined group of old Swedish cultivars might have an ancestry that differs from that of the new Swedish cultivars and from most of the foreign cultivars.

Several different genes have been proposed as responsible for fruit texture variability at harvest and/or after storage. We have analysed 127 apple cultivars for allelic composition in two key genes that are directly involved in the ethylene biosynthesis pathway, Md-ACS1 and Md-ACO1, and two other genes that are involved in cell wall degradation, Md-Exp7 and Md-PG1. Firmness was measured with a penetrometer at harvest and after 6 or 12 weeks (early- and late-maturing cultivars, respectively) of cold storage. Maturation time was positively correlated with firmness at harvest and negatively correlated with fruit softening rate (difference between firmness at harvest and after storage, divided by number of weeks in storage). Polyploid cultivars showed significantly higher firmness at harvest compared to diploids. Alleles previously described as responsible for good texture were associated with significantly lower softening for Md-ACS1 and Md-PG1, but the opposite was noted for Md-EXP7. Results were nonsignificant for Md-ACO1. Allele frequencies were very uneven in all four loci, with the three most common multi-locus configurations accounting for 64 % of the entire material. The predictive power of these genes was calculated with a partial least squares discriminant analysis, and these accounted for 15 % of the observed variation in initial firmness and 18 % for softening rate. Inclusion of maturation time, storage time (i.e. 6 or 12 weeks) and initial firmness into the model however increased the predictability of softening rate to 38 %. Dividing the material in modern (released after 1960) and old cultivars did not change the outcome of our analyses.

Delta-like 1 homolog (Dlk1) and insulin-like growth factor 2 (Igf2) are two of six well-studied mouse imprinted gene clusters that are paternally expressed. Their expression is also linked to their maternally expressed non-coding RNAs, encoded by Gene trap locus 2 (Gtl2) and Imprinted maternally expressed transcript (H19), co-located as imprinted gene clusters. Using deoxyoligonucleotide microarrays and real-time RT-PCR analysis we showed Dlk1 and Gtl2 to exhibit a time-course of expression during tooth development that was similar to that of Igf2 and H19. Western blot analysis of proteins encoded by Dlk1 and Igf2 suggested that the levels of these proteins reflected those of the corresponding mRNAs. Immunohistochemical studies of DLK1 in murine molars detected the protein in both epithelial and mesenchymal regions, in developing cusp mesenchyme, and in newly synthesized enamel and dentin tubules. IGF2 protein was detected primarily at prenatal stages, suggesting that it may be active before birth. Analysis of methylation of cytosine-phosphate-guanine (CpG) islands in both Dlk1 and Igf2 suggested the presence of an increasing fraction of hypermethylated bases with increasing time of development. The increased levels of hypermethylation coincided both with the diminished levels of expression of Dlk1 and Igf2 and with decreased levels of DLK1 and IGF2 proteins in the tooth germ, suggesting that their expression is regulated via methylation of CpG islands present in these genes.

Several different genes have been reported to affect fruit texture in apple. We have analysed 158 cultivars for their allelic composition in the expansin gene MdExp7. Most of these cultivars had already been analysed for their allelic composition in the ethylene production-affecting gene Md-ACS1 but 29 additional cultivars were analysed also for this gene. Finally, we harvested fruit from 114 cultivars and measured firmness with a penetrometer directly at harvest and after 6 weeks (earlyripening) or 12 weeks (late-ripening cultivars) of cold storage. For the Md-Exp7 locus, an SSR allele with 198 bp has previously been suggested as a marker for the ability to retain fruit firmness during storage. However, we found that another marker with 202 bp was over-represented in (1) cultivars that also carry the favourable allele 2 in the Md-ACS1 locus, and in (2) more recently developed cultivars, both of which suggest that recent selection in several apple breeding programs around the world has favoured allele 202 more than allele 198. In addition, preliminary data from the firmness evaluations also suggest that allele 202 is more closely associated with higher initial firmness and better firmness retention during storage. Firmness was retained during storage to a very high degree by two ‘Antonovka’ types in spite of their unfavourable alleles in the Md-ACS1 and Md-Exp7 loci. These cultivars could thus be valuable in the search for additional fruit texture genes.

BACKGROUND: Wilms tumor (WT) is the most common renal neoplasm in children. Histologically, most WTs consist of three tissue elements: blastema, epithelium, and stroma. Some cases also show diffuse or focal anaplastic features. Previous studies have shown that a predominance of blastemal cells in post-chemotherapy WT specimens is associated with a poor clinical course. However, there is currently no molecular marker for blastemal cells, and risk stratification for post-nephrectomy treatment is therefore often based on clinico-histological parameters alone. PROCEDURE: In the present study, three public gene expression microarray datasets, including 82 WTs and 8 normal fetal kidneys, were used to establish a consensus gene expression profile of WT. By bioinformatic analyses, 17 genes overexpressed in WT compared to fetal kidney were then selected for evaluation of their protein expression in WT cell lines and in the different histological components in paraffin-embedded WT tissue sections by immunofluorescence. RESULTS: Most of the evaluated proteins were expressed in all three common histological components. A prominent exception was SIX1, being expressed predominantly in blastemal elements in 24/25 pediatric cases containing blastema. Anaplastic elements exhibited highly variable SIX1-positivity. The SIX2 protein, known to be co-expressed with SIX1 during nephrogenesis, only exhibited blastemal-predominant expression in half of the SIX2 evaluated cases. CONCLUSIONS: Genes highly expressed in WT compared to fetal kidney are generally overexpressed in all of the three common WT tissue elements. An exception is the predominant expression of SIX1 in blastemal cells, hereby identifying this protein as a candidate marker for blastema. Pediatr Blood Cancer © 2011 Wiley Periodicals, Inc. (Less)

Fire blight (Erwinia amylovora) causes serious damage to pome fruit orchards, and identification of germplasm with heritable disease resistance is therefore crucial. Two dominant SCAR (sequence characterised amplified region) marker alleles (AE10-375 and GE-8019), flanking a previously identified QTL (quantitative trait locus) for resistance to fire blight on ‘Fiesta’ linkage group 7 in apple cultivars related to ‘Cox’s Orange Pippin’, were screened on 205 apple cultivars. Both marker alleles were present in 22% of the cultivars, indicating presence of the QTL allele for tolerance, and both were lacking in 25%, indicating homozygosity for absence of the QTL tolerance allele. However, 33% had only the marker allele AE10-375, while 20% had only GE-8019, suggesting that some cultivars with the dominant alleles for both of the flanking markers can carry these on separate chromosomes and may lack the QTL allele for tolerance. In 2009 and 2010, terminal shoots of greenhouse-grown grafted trees of 21 cultivars (only 20 in 2010) were inoculated with Erwinia amylovora. ‘Idared’ (susceptible) and ‘Enterprise’ (tolerant) were included as controls. Disease severity for each cultivar was expressed as percentage of necrosis in relation to entire length of shoot, and the ranking of cultivars in 2009 and 2010 was compared with a Spearman rank correlation test, P

In order to gain insight into possible cellular functions of the prion protein (PrP) during normal development, the expression of Prnp (encoding the PrP) and the distribution of the PrP were studied in murine tooth germs. Expression of Prnp in the mouse first molar tooth germ was highly dynamic, increasing several-fold during the secretory phase of odontogenesis, exhibiting a time-course of expression similar to that of genes coding for other extracellular proteins [e.g. enamel matrix proteins (Amelx, Ambn, Enam), Aplp1, Clstn1, and Clu]. Western blot analysis suggested that the amounts of PrP and amyloid beta (A4) precursor-like protein 1 (APLP1) in the tooth germ followed time-courses similar to those of the corresponding mRNAs. Immunohistochemical studies of the distribution of PrP in murine molar and incisor tooth germs at embryonic day (E)18.5 suggested that this protein was located in the cervical loop, outer enamel epithelium, pre-ameloblasts, and dental papilla. Different degrees of immunolabelling of pre-ameloblasts on the mesial and distal aspects of a lower molar cusp may be related to different enamel configurations on the two aspects. It is concluded that the dynamic patterns of expression of Prnp, and of distribution of PrP, suggest that PrP may have functions during secretory odontogenesis, perhaps in relation to amelogenesis.

Low polymorphism in cultivated watermelon has been reported in previous studies, based mainly on US Plant Introductions and watermelon cultivars, most of which were linked to breeding programmes associated with disease resistance. Since germplasm sampled in a putative centre of origin in southern Africa may harbour considerably higher variability, DNA marker-based diversity was estimated among 81 seedlings from eight accessions of watermelon collected in Zimbabwe; five accessions of cow-melons (Citrullus lanatus var. citroides) and three of sweet watermelons (C. lanatus var. lanatus). Two molecular marker methods were used, random amplified polymorphic DNA (RAPD) and simple sequence repeats (SSR) also known as microsatellite DNA. Ten RAPD primers produced 138 markers of which 122 were polymorphic. Nine SSR primer pairs detected a total of 43 alleles with an average of 4.8 alleles per locus. The polymorphic information content (PIC) ranged from 0.47 to 0.77 for the RAPD primers and from 0.39 to 0.97 for the SSR loci. Similarity matrices obtained with SSR and RAPD, respectively, were highly correlated but only RAPD was able to provide each sample with an individual-specific DNA profile. Dendrograms and multidimensional scaling (MDS) produced two major clusters; one with the five cow-melon accessions and the other with the three sweet watermelon accessions. One of the most variable cow-melon accessions took an intermediate position in the MDS analysis, indicating the occurrence of gene flow between the two subspecies. Analysis of molecular variation (AMOVA) attributed most of the variability to within-accessions, and contrary to previous reports, sweet watermelon accessions apparently contain diversity of the same magnitude as the cow-melons.

At the mouse incisor tip the initially differentiated ameloblasts produce a thin, prism-free enamel, while further apically, in the immediate adjacent segment, the enamel thickness increases and the four-layered enamel of mouse incisor is formed. Comparative gene-expression profiling was carried out on RNA isolated from these two segments of incisor tooth germs at embryonic day (E)17.5 and at postnatal days (P)0, 1, 2, and 10 using microarrays to measure messenger RNA (mRNA) and microRNA (miRNA) species present in the segments. Validation of expression data was achieved using real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blotting. Bioinformatic data suggested enhanced cellular apoptosis in the incisal tip segment, which, together with diminished expression of the Amelx and Enam genes, may contribute to the production of the thin enamel seen in this tooth segment. For genes exhibiting higher levels of expression in the adjacent segment where complex enamel is being formed, bioinformatic analysis suggested significant associations with cellular functions involving the actin cytoskeleton, cellular development, morphology, and movement. This is suggested to reflect that ameloblasts with Tomes' process are being organized in transverse rows, facilitating the transverse movement that results in prism decussation in the inner enamel of the adjacent segment. Bioinformatic analysis of miRNA expression data lends support to these suggestions.

Trees of 68 apple cultivars, aimed for preservation by the ‘National Program for diversity of cultivated plants’ as mandate cultivars, were analysed using a set of 10 SSR (simple sequence repeat) primer pairs and the self-incompatibility (S � )locus to evaluate genetic diversity and reveal inter-cultivar relationships. The 12 polymorphic SSR loci exhibited 2 to 15 alleles, with expected heterozygozity (He) ranging from 0.36 to 0.88 and a mean of 0.74. Numerous alleles were classified as rare or unique (35% and 18% respectively). For the S-locus, a total of 14 alleles were identified in this study. Five alleles, S1� S3 ,S 5 and S7 had frequencies ranging from 11 to 18%, whereas the remaining 9 alleles were below 6%. All sexually obtained cultivars could be distinguished with the set of SSR loci. Sports were identical with their progenitors in two cases, but differed in one SSR allele in a third case. An SSR-based dendrogram, based on Roger’s genetic distances, did not reveal any clear pattern of clustering. The genetic distances were, however, correlated with a corresponding matrix obtained in a previously conducted RAPD-based study of the same cultivars. Non-mandate parents of Swedish mandate cultivars together with some other reference cultivars were included in this study to check the accuracy of allele scoring, verify parentage and compare the results of this study with those presented in previously published studies. Some discrepancies in allele sizing were revealed and the possibilities of avoiding this problem are discussed.

SummaryGametic self-incompatibility in apple is determined by a set of S-locus alleles, which can be identified using PCR with allele-specific DNA primers. Allele composition has previously been determined for a large number of apple cultivars grown in North America, central and southern Europe, and Asia. In the present study, 103 not previously studied apple cultivars that are grown mainly in northern Europe were scored for their S-allele composition in order to provide apple growers and breeders with information on incompatibility among cultivars. In addition, S-allele frequencies were determined in some control cultivars and compared with data reported previously. A different genotype was found for ‘Discovery’, which was therefore added to our list. The most common S-allele in the resulting set of 104 cultivars investigated at Balsgard was S7 (18%) followed by S3 (17%), S5 (14%), and S1 and S2 (both at 11%). Comparisons of allele frequencies obtained from previous compilations showed that the frequency...

SummaryInreasing consumer demand for firm apple fruit has raised interest among scientists and breeders to identify heritable traits that affect fruit texture. One such candidate gene is the bi-allelic Md-ACS1 locus, that shows a strong association between genotype and internal ethylene content in ripening fruit. However, associations between genotype and fruit firmness, or between genotype and fruit softening during storage, have been somewhat unclear. In order to determine the potential importance of allelic variation at this locus, we used PCR methods to investigate Md-ACS1 alleles in 137 apple cultivars. In addition, we also compiled data from previously published studies, to achieve an eventual dataset with 270 cultivars. Allele 1, associated with higher ethylene concentrations in ripening apples, was much more common in older cultivars (relative frequency = 0.88 in cultivars introduced before 1800; 0.84 in cultivars introduced between 1800 – 1899; and 0.70 in cultivars introduced in 1900 – 1959). Co...

Ameloblastomas are a class of odontogenic tumors, which arise from developmental remnants in the oral tissue. Although the cellular and molecular mechanisms resulting in development of ameloblastoma are poorly understood, it is generally accepted that they exhibit an odontogenic source and originate from epithelial cells associated with tooth development. The epithelial sources in the oral tissue that can cause ameloblastomas include enamel organ, reduced enamel epithelium, rests of Malassez, and rests of Serres. These remnants originate from the stomodeal ectoderm, which give rise to the oral epithelium that initiate and guide tooth development as the embryo develops. It is of great clinical value to understand the developmental origin of these epithelial components and their histology, since the ameloblastomas display histopathological similarities to their structures.

Indications exist in the scientific literature that infection with human herpes family viruses may contribute to the pathogenesis of neuroblastoma (NB). However, systematic investigations regarding viral presence in NB cells have been scarcely reported. Here, the presence of DNA from Epstein-Barr virus (EBV) and human cytomegalovirus (HCMV) was assessed by PCR in 12 NBs, supplemented with RNA in situ hybridization, immunohistochemical detection, and high-throughput DNA sequencing. These standard methods did not detect infection by EBV or HCMV in NB cells in any tumor, while occasional immune cells were positive for EBV RNA or HCMV protein in four cases.

Conclusions A clear effect on gene expression in the developing tooth germ was apparent after 24 h at all dosages. The results indicate increased transcription of genes involved in development of bone, increased biosynthesis of important carbohydrates and of enamel/dentin, and reduced expression of apoptosis related proteins. Further investigations are, however, required to elucidate these findings. Such experiments will likely entail the establishment of a clear dose-response relationship and long term use of MK-7. Also, effects of oral administration should be studied. Background Menaquinone-7 (MK-7) it’s a fat-soluble vitamin and a vitamin K2 homologe needed for post-translational modification of certain proteins required for blood coagulation, and in metabolic pathways in bone and other tissues. Recent studies found an association between long-term anticoagulant treatment (OAC) and reduced bone quality due to reduction of active osteocalcin. OAC is often linked to an undesired soft-tissue calcification in both children and adults. OAC often leads to increased incidence of fractures, reduced bone mineral density/bone mineral content, osteopenia and increased serum levels of undercarboxylated vitamin Kdependent proteins, known as Gla-proteins. Litle is known about the effects of vitamin K2 during tooth development.

Expression of clusterin (Clu) in the murine first molar tooth germ was markedly increased at postnatal developmental stages. The time-course of expression of this gene paralleled those of other genes encoding proteins involved during the secretory phase of odontogenesis, as described previously. Immunohistochemical studies of clusterin in murine molar tooth germs suggested this protein to be located in outer enamel epithelium, regressing enamel organ, secretory ameloblasts, and the dental epithelium connecting the tooth to the oral epithelium at an early eruptive stage. Immunolabelling of transforming growth factor beta-1 (TGF-β1) revealed it to be located close to clusterin. The levels of expression of Clu and Tgfb1 were markedly decreased following in-vivo transfection with anti-miR-214. In contrast, the expression of several genes associated with regulation of growth and development were increased by this treatment. We suggest that clusterin has functions during secretory odontogenesis and the early eruptive phase. Bioinformatic analysis after treatment with anti-miR-214 suggested that, whilst cellular activities associated with tooth mineralization and eruption were inhibited, activities associated with an alternative developmental activity (i.e. biosynthesis of contractile proteins) appeared to be stimulated. These changes probably occur through regulation mediated by a common cluster of transcription factors and support suggestions that microRNAs (miRNAs) are highly significant as regulators of differentiation during odontogenesis.

European canker, caused by the fungus Neonectria ditissima, is a severe problem in apple production both in Sweden and in many other northern European countries. Even when applying fungicides and good horticultural practices, canker damage occurs almost yearly in nurseries and orchards. Some years, devastating outbreaks destroy numerous trees. To date, complete resistance to N. ditissima is not known in apple. For further research and plant breeding, heritable variation in quantitative resistance should be investigated by phenotyping large sets of cultivars. In the present project, 55 apple cultivars were screened for resistance to N. ditissima. One-year-old shoots from mature trees were inoculated in the greenhouse with a standardized volume and concentration of conidia suspension using different inoculation methods. Two-year-old trees of five cultivars were inoculated in the field. Length of the occurring cankers was measured at regular intervals throughout a period of up to three months. The investigated cultivars showed considerable differences in colonization rate. In cultivars known to be highly resistant, i.e., ‘Santana’, lesions progressed much slower compared to susceptible cultivars like ‘Cox’s Orange Pippin’ and ‘James Grieve’. Since the inoculation-based phenotyping is demanding in labour and time (duration), especially when the test is performed on grafted trees, qPCR-based assessment of fungal biomass at early stages of infection was explored as an alternative or complementary approach for phenotyping.

Neuroblastomas (NBs) are tumours of the sympathetic nervous system accounting for 8-10% of paediatric cancers. NBs exhibit extensive intertumour genetic heterogeneity, but their extent of intratumour genetic diversity has remained unexplored. We aimed to assess intratumour genetic variation in NBs with a focus on whole chromosome changes and their underlying mechanism. Allelic ratios obtained by SNP-array data from 30 aneuploid primary NBs and NB cell lines were used to quantify the size of clones harbouring specific genomic imbalances. In 13 cases, this was supplemented by fluorescence in situ hybridisation to assess copy number diversity in detail. Computer simulations of different mitotic segregation errors, single cell cloning, analysis of mitotic figures, and time lapse imaging of dividing NB cells were used to infer the most likely mechanism behind intratumour variation in chromosome number. Combined SNP array and FISH analyses showed that all cases exhibited higher inter-cellular copy number variation than non-neoplastic control tissue, with up to 75% of tumour cells showing non-modal chromosome copy numbers. Comparisons of copy number profiles, resulting from simulations of different segregation errors to genomic profiles of 120 NBs indicated that loss of chromosomes from a tetraploid state was more likely than other mechanisms to explain numerical aberrations in NB. This was supported by a high frequency of lagging chromosomes at anaphase and polyploidisation events in growing NB cells. The dynamic nature of numerical aberrations was corroborated further by detecting substantial copy number diversity in cell populations grown from single NB cells. We conclude that aneuploid NBs typically show extensive intratumour chromosome copy number diversity, and that this phenomenon is most likely explained by continuous loss of chromosomes from a polyploid state.

It is generally believed that fever is mediated by certain cytokines produced by immune cells activated by exogenous pyrogens, e.g., lipopolysaccharides (LPS), released into the circulation and transported to the brain. There, the cytokines are thought to stimulate prostaglandin (PG) E 2 production within the organum vasculosum laminae terminalis region. PGE, then may act as a febrigenic mediator locally or in the surrounding preoptic area (POA). However, whereas the increases in preoptic PGE 2 and body (core) temperature (T c ) following the intravenous (i.0 administration of LPS correlate temporally, cytokine levels in blood lag both these increases. From recent data in the literature, we have conjectured that a possible, alternative communication pathway between the i.v. LPS-activated immune system and brain PGE 2 may be provided by the vagi. To test this possibility, we measured the levels of PGE 2 in the extracellular fluid of the POA (collected by microdialysis) of conscious, subdiaphragmatically vagotomized or sham-operated guinea pigs following LPS administration (2 μg/kg; i.v.); controls received pyrogen-free saline (PFS). The effluents from the microdialysis probes were collected over 30-min periods throughout the experiments and the samples analyzed by radioimmunoassay; (T c ) was monitored continuously using thermocouples inserted 5 cm into the colon. LPS induced a biphasic fall in T c and failed to increase preoptic PGE 2 levels in the vagotomized guinea pigs ( n = 10 ), whereas in their sham-operated controls ( n = 10 ) it induced increases in both preoptic PGE 2 and (T c ) within 15 min after its injection; PFS ( n = 13 ) had no effect on either variable. We postulate that peripheral immune cell-derived signals may be transmitted via the vagi to the medulla. From other data, we suggest further that they may be conveyed from here via the ventral noradrenergic bundle to the POA region, where the released norepinephrine induces the local synthesis of PGE 2 and, hence, fever onset.