Bone morphogenetic proteins (BMPs) are multifunctional secreted growth factors regulating a broad spectrum of functions in numerous systems. An increased expression and production of specific BMPs have been described in the rheumatoid arthritis (RA) synovium. The aim of this study was to analyze the involvement of the BMP signaling pathway in RA synoviocytes in response to interleukin-17 (IL-17) and tumor necrosis factor-alpha (TNF-α).The expression of components of the BMP signaling pathway  was analyzed by quantitative polymerase chain reaction before and after treatment of RA synoviocytes with TNF-α or IL-17 or both. Regulation was studied in the presence of the specific BMP inhibitor DMH1 (dorsomorphin homologue 1) or an exogenous BMP ligand, BMP6. Expression and production of pro-inflammatory cytokines (IL-6 and granulocyte-macrophage colony-stimulating factor), chemokines , and matrix metalloproteinases  were analyzed.RA synoviocytes express BMP receptors , signal transducers of the Smad family (Smad1 and 5 and co-Smad4), and different BMP antagonists. The modulation of the expression of the BMP target genes—Id (inhibitor of DNA-binding/differentiation) proteins and Runx (Runt-related transcription factor) transcription factors—after the addition of exogenous BMP shows that the BMP signaling pathway is active. RA synoviocytes also express BMP ligands  which are highly upregulated after activation with TNF-α and IL-17. Autocrine BMP signaling pathway can be blocked by treatment with the inhibitor DMH1, leading to an increase in the upregulated expression of pro-inflammatory cytokines, chemokines, and MMPs induced by the activation of RA synoviocytes with TNF-α and IL-17. Conversely, the additional stimulation of the BMP pathway with the exogenous addition of the BMP6 ligand decreases the expression of those pro-inflammatory and pro-destructive factors.The results indicate that the canonical BMP pathway is functionally active in human RA synoviocytes and that the inhibition of autocrine BMP signaling exacerbates the pro-inflammatory phenotype induced in RA synoviocytes by the stimulation with IL-17 and TNF-α. Bone morphogenetic proteins (BMPs) are secreted signaling proteins which form a subgroup of the transforming growth factor-beta (TGF-β) superfamily . BMPs arBMPs were originally identified as growth and differentiation factors for osteogenic cells but now are considered multifunctional proteins implicated in the development of virtually all organs and the renewal and maintenance of different adult tissues , 4–6. ThDespite all these data, however, only a few studies have addressed the involvement of BMP signaling in RA, pointing out a role for BMP6 and BMP7 in stimulating the survival and the proliferation and extracellular matrix component biosynthesis, respectively, in synoviocytes , 14. In Synoviocytes were grown from synovial tissue samples obtained from RA patients undergoing joint surgery. The patients with RA fulfilled the American College of Rheumatology criteria for RA . Each in4 cells/cm2. Cells then were treated for 12 h with different combinations of IL-17 50 ng/mL , TNF-α 0.5 ng/mL (R&D Systems), BMP-6 10 ng/mL , and dorsomorphin homologue 1 (DMH1) 30 μM . After 12-h treatment, RNA was extracted by using an RNeasy Plus Mini Kit  in accordance with the instructions of the manufacturer. RNA then was quantified by using a Qubit RNA BR Assay Kit (Life Technologies).Cells were seeded in 12-well plates at a density of 5×10Total cDNA was synthesized by using a High Capacity cDNA Reverse Transcription Kit  in accordance with the instructions of the supplier and then used as a target in the polymerase chain reaction (PCR) amplifications. Pre-designed TaqMan gene expression assays for different genes were obtained from Applied Biosystems. GNB2L1 was used as an endogenous control. All PCRs were set in duplicates by using the TaqMan Gene Expression Master Mix (Applied Biosystems) in accordance with the instructions of the manufacturer. Amplifications, detections, and analyses were performed in a 7.900HT Fast Real-time PCR System . The ∆Ct method was used for normalization to GNB2L1.Culture supernatants of RA synoviocytes were harvested after 12 h of treatment. IL-8 secretion was measured by using an enzyme-linked immunosorbent assay  and IL-6 and granulocyte-macrophage colony-stimulating factor (GM-CSF) were measured by using Cytometric Bead Array assays  in accordance with the instructions of the manufacturers.P ≤ 0.05 (*), P ≤ 0.01 (**), and P ≤ 0.001 (***) were considered to be statistically significant.The Mann–Whitney test was used to compare differences. Values of The expression of different components of the BMP signaling pathway was analyzed in RA synoviocytes. BMP receptor complexes are constituted by one of the following type I receptors, type IA BMP receptor (BMPRIA)/ALK-3, type IB BMP receptor (BMPRIB)/ALK-6, and type IA activin receptor (ACTRIA)/ALK-2, which commonly combine with type II BMP receptor (BMPRII) but also the type IIA and IIB activin receptors (ACTRIIA and ACTRIIB). In RA synoviocytes, BMPRIA and ACTRIA were the most abundantly expressed type I receptors, and the BMP-specific type II receptor BMPRII showed a high level of expression Fig. .Fig. 1ExAmong ligands, the most highly expressed were BMP2 and BMP6 Fig. . SynovioFurthermore, BMP signaling is finely regulated at multiple levels: at the extracellular space, several high-affinity antagonists bind selectively with BMPs to inhibit their biological actions; at the plasma membrane, the pseudoreceptor BAMBI associates with BMP receptors, preventing the formation of active receptor complexes; and intracellularly, inhibitory Smads and Smurfs inhibit further signaling and activation of target genes . The expThe functionality of the BMP pathway was analyzed by studying the modulation of the expression of the components of Id protein and Runx families after addition of exogenous BMP. As shown in Fig. We next examined the effects of critical pro-inflammatory cytokines in RA, such as TNF-α and IL-17, on the expression of the different components of the BMP signaling pathway. The culture of synoviocytes with IL-17 and TNF-α alone or in combination induced mainly a 3- to 4-fold increase in the expression of BMPRIB, the least expressed type I BMP receptor in unstimulated synoviocytes, and slight increases in the mRNA levels of BMPRII and ACTRIIB receptors Fig. . The expThe above data showing BMP receptor expression on synoviocytes, along with an upregulated BMP expression after pro-inflammatory cytokine stimulation, suggested that BMP ligands produced by RA synoviocytes could act in an autocrine manner. The highly selective type I BMP receptor inhibitor DMH1, which blocks the canonical BMP signaling pathway by inhibiting the BMP-induced Smad 1/5/8 activation , was useThe expression of chemokines other than IL-8 was analyzed, and the results showed that BMP signaling inhibition mainly augmented the expression of CCL2 induced by IL-17 Fig. . HoweverThe treatment of RA synoviocytes with IL-17 or TNF-α or both also upregulated the expression of MMP-2 and MMP-3 metalloproteinases involved in synoviocyte migration and invasion as well as joint destruction , 23 Fig. When thThe effects of the addition of an exogenous BMP ligand, inducing an additional BMP signaling pathway activation, were also addressed. BMP6 was chosen because of its notable expression levels in RA synoviocytes and its high upregulation after synoviocyte stimulation with pro-inflammatory cytokines Figs.  and 3b. The increased expression and production of specific BMPs  in the synovium and synovial fluid of patients with RA as well as in mouse models of arthritis have been reported by different groups –15. HoweIn this study, we show the presence of a functionally active BMP signaling pathway in synoviocytes. Human RA synoviocytes express the three type I BMP receptors, BMPRIA, BMPRIB, and ACTRIA, and the BMP-specific type II receptor BMPRII. RA synoviocytes also express the BR-Smads as well as the common partner Smad4. All of these components are found to be efficient in signal transduction given the ability of synoviocytes to modulate the expression of some BMP target genes, including some Runx factors and members of the Id protein family, in response to exogenous BMP stimulation. In addition, RA synoviocytes express BMP ligands which, in agreement with previous work , 18, 19,The blockade of BMP signaling with DMH1 significantly enhances the expression of pro-inflammatory cytokines , chemokines , and MMPs  induced in RA synoviocytes by the simultaneous treatment with IL-17 and TNF-α alone or in combination. Conversely, the stimulation of the BMP signaling pathway with an exogenous BMP ligand reduces the expression of pro-inflammatory and pro-destructive factors in both unstimulated and stimulated RA synoviocytes, mainly with the IL-17 and TNF-α combination. Therefore, it could be hypothesized that BMP signaling could have an anti-inflammatory role in the control and maintenance of low levels of pro-inflammatory factors in healthy synoviocytes and probably also in the early stages of RA (Fig. However, the stimulation of RA synoviocytes induces the expression not only of BMP ligands but also of BMP antagonists, mainly after treatment with TNF-α alone or in combination with IL-17. Smad7 and BAMBI are the main upregulated BMP antagonists, which act at the intracellular and the plasma membrane level, respectively , indicatIn this context, the high levels of BMP ligands produced by stimulated synoviocytes could also contribute to the chronic inflammation associated with RA by acting in a paracrine fashion on surrounding cells present in the inflamed synovium Fig. . BMPs haWe propose that BMPs would play a dual role in the synovium, controlling the production of pro-inflammatory cytokines and inhibiting synoviocyte transformation in healthy and non-inflammatory conditions, but in advanced stages of RA could contribute to chronic inflammation, allowing the functional deregulation of synoviocytes and promoting the activation and pro-inflammatory functions of immune and non-immune cells accumulating in the inflamed arthritic joint.


The aim of the present study was to investigate the significance of survivin protein expression levels in craniopharyngioma. Tumor samples and clinical data were obtained from 50 patients with craniopharyngioma who were admitted to the West China Hospital of Sichuan University . The morphology of the craniopharyngioma samples was observed using optical and electron microscopes, and survivin expression was investigated in the samples by immunohistochemical analysis. The immunohistochemical results revealed survivin expression in all of the craniopharyngioma samples, but not in the healthy brain tissue samples. It was identified that survivin was expressed at a higher level in cases of the adamantinomatous type compared with those of the squamous-papillary type, in male patients compared with female patients, in children compared with adults and in recurrent cases compared with non-recurrent cases. Furthermore, no significant difference was detected in survivin expression levels among the tumors of different subtypes and different disease stages. The results of the present study indicate that survivin is significant in the development of craniopharyngioma, and that survivin protein expression levels are a meaningful indicator for assessing craniopharyngioma recurrence. Craniopharyngioma is a slow-growing benign tumor that develops in the parasellar or sellar region of the brain ,2. AccouDespite continuous progress in microsurgery techniques and years of research, the treatment of craniopharyngioma remains a significant problem, particularly for children . The uniIn the present study, the ultrastructure of craniopharyngioma was observed using electron microscopy and immunohistochemistry, with the aid of digital image analysis techniques, and survivin protein expression levels in craniopharyngioma were assessed to determine the biological characteristics and pathogenesis of craniopharyngioma.The paraffin-embedded samples of 50 patients admitted to the West China Hospital of Sichuan University  and diagnosed with craniopharyngioma following surgical resection between January 2000 and December 2005 were selected for use in the present study. The inclusion criteria were as follows: i) A complete patient medical history must be available; ii) the patient must have undergone pre-operative imaging (computed tomography or magnetic resonance imaging); iii) the pathology sample must be a sufficient size and have been established as craniopharyngioma by the Department of Pathology of the West China Hospital; and iv) the patient must have not have received radiation therapy, so as to eliminate the impact of radiation. According to the standard classification of central nervous system tumors published by the World Health Organization in 2000 , 33/50 cSurvivin protein expression levels in the craniopharyngioma tissues were detected using immunohistochemistry and then quantitatively analyzed. Apoptotic cells were stained with transferase dUTP nick end labeling-peroxidase  and observed using transmission electron microscopy . Under the light microscope, survivin-positive cells were observed as brown granules in the nucleus or cytoplasm. For each section, a charge-coupled device camera was used to capture at least five fields of view at a magnification of ×400 , and Image-Pro Plus software  was used to quantitatively analyze the images; integrated optical density (IOD) was selected as the parameter to indicate survivin protein expression levels and the average IOD of all the recorded images was determined. Furthermore, TUNEL-POD stained the nuclei of the apoptotic cells yellow or brown, while the nuclei of the non-apoptotic cells were blue.Statistical analysis was performed using SPSS software, version 13.0 . Non-parametric tests were performed for each factor, and multinomial and ordinal logistic regression was performed. P<0.05 was used to indicate a statistically significant difference.Adamantinomatous  and squaDifferent types of tumor cells were observed using electron microscopy. The nuclei of the apoptotic tumor cells  were ovaSurvivin protein expression was detected in all 50 craniopharyngioma samples in the cytoplasm or the nucleus. In adamantinomatous craniopharyngioma , signifiThe images of et al ; patients with no complications exhibited a significantly more favorable prognosis compared with patients who experienced complications. However, no significant correlation was detected among all other candidate risk factors.Controversy regarding the origin of craniopharyngioma has always been present, and two expounding theories currently exist: The embryonic origin theory and the metaplasia theory. Certain individuals believe that it is the residual ectoderm cells in the primary cranial pharyngeal tube and anterior pituitary that form craniopharyngioma ,19. In tet al (et al (et al (Histologically, craniopharyngioma can be divided into the adamantinomatous and squamous-papillary types. The majority of children with craniopharyngioma exhibit the adamantinomatous form, however, an equal proportion of the two types are observed in adults . Furtheret al  identifil (et al  identifil (et al  demonstret al (In 1997, survivin, a novel inhibitor of apoptosis, was identified . A high et al  evaluateIn conclusion, the current study indicates that survivin may regulate the cell cycle and apoptosis in the tumorigenesis of craniopharyngioma. Survivin may significantly inhibit the apoptosis of the tumor cells, thus increasing the aggressiveness of benign tumors and the malignancy of malignant tumors, as well as increasing the likelihood of tumor recurrence. Therefore, survivin may be useful as a predictor of craniopharyngioma patient prognosis, however, additional studies on larger patient samples are required to confirm this association.


MCT) is a new, open-access journal, devoted to molecular mechanisms, preclinical and clinical research and development of gene-, peptide-, protein-, and cell-based therapies.Gene therapy is suggested to be one of the most specific and efficient modulations for gene deficient diseases and extended to other diseases like cancer and inflammation, even though there are still challenges to be faced, such as specific and selective delivery, minimal to no toxicity, efficient metabolism, simplicity, and measurable efficiency. It is important to identify and validate drug-able disease-specific targets for molecular and cellular therapies, while it is equally important to have disease biomarkers to trace and define the biological effects of molecular and cellular therapies. The importance and significance of allostery in molecular and cellular therapies and “allosteric disease”, “allosteric effect”, and “allosteric drug” should be more carefully examined and validated. Cell therapy has been attracting an increasing amount of consideration in the development of new treatments for diseases. Molecular and Cellular Therapies ( MCT), a new international journal devoted to molecular mechanisms, preclinical and clinical research and development of gene-, peptide-, protein-, and cell-based therapies, including target identification and validation, safety, toxicity, efficacy, efficiency, pharmacokinetics, metabolism, formulation, delivery, pharmacovigilance, biomarkers, and new drug applications. MCT will integrate interdisciplinary fields such as genetics, cell biology, drug development, therapeutics, and new methods in biotechnology toward translational medicine. MCT aims to establish a scientific channel to translate the understanding of molecular and cellular therapies to clinical and medical applications, to improve the survival rate and quality of life for patients.Welcome to the open-access journal titled Molecular and Cellular Therapies  is a key component of the stem cell niche that can regulate stem cell behaviour, renewal or differentiation. Within the ECM there are different types of protein interactions that can trigger various cell responses via diverse sensing mechanisms and downstream signaling pathways . The intProtein-based drug development has been experiencing a vast array of opportunities as well as several challenges to overcome, even though it is well accepted that protein-based drugs have higher affinity and specificity from protein-protein interactions and are more effective to target and modulate the signaling pathways responsible for the occurrence of diseases. Brentuximab vedotin (SGN-35) is an anti-CD30 monoclonal antibody (cAC10) conjugated by a protease-cleavable linker to a microtuxbule-disrupting agent, monomethyl auristatin E. Brentuximab vedotin, has been approved as an effective treatment for relapsed CD30-expressing Classical Hodgkin and systemic anaplastic large cell lymphomas. Brentuximab vedotin was found to decrease cell proliferation, induce cell cycle arrest, trigger apoptosis of primary effusion lymphoma cells, inhibit tumor growth, and improve the survival of animals with primary effusion lymphoma . BrentuxMCT is the journal focusing on the clinical application of gene-, peptide-, protein-, and cell-based drugs and keeping track of the wealth of new information related to the topic. We have the confidence to believe that the MCT will play an important, critical, and recognized role in understanding the molecular mechanisms of molecular and cellular therapies and developing the individual medicine and therapeutic strategy.There is an urgent and immediate need to create a forum to stimulate discussion and exchange of scientific findings and understandings of molecular and cellular therapies with a clear goal of treating diseases and improving the quality of patients. MCT, are delighted to welcome you to this new and exciting journal and thank the scientists who have agreed to publish in the journal. In setting up the journal, we owe an enormous debt of gratitude to all professors and scientists for their encouragement, support, comments, suggestions, and contributions. With great supports from our Section Editors, Associate Editors and Editorial Board Members, we deeply believe that MCT will be well-received both by preclinical, clinical and pharmaceutical scientists interested in molecular and cellular therapies and contribute to better outcome for understanding the diseases and developing new therapies. Involvement and contributions from a large group of scientists who work on molecular and cellular therapies are crucial to the success of the journal.In conclusion, we as Editors of


Lanice conchilega . The aggregations are known to have a considerable impact on the physical and biogeochemical characteristics of their environment and subsequently on the abundance and biomass of primary food sources and the macrofaunal  community. Therefore, we hypothesise that L. conchilega aggregations affect the structure, stability and isotopic niche of the consumer assemblage of a soft-bottom intertidal food web. Primary food sources and the bentho-pelagic consumer assemblage of a L. conchilega aggregation and a control area were sampled on two soft-bottom intertidal areas along the French coast and analysed for their stable isotopes. Despite the structural impacts of the ecosystem engineer on the associated macrofaunal community, the presence of L. conchilega aggregations only has a minor effect on the food web structure of soft-bottom intertidal areas. The isotopic niche width of the consumer communities of the L. conchilega aggregations and control areas are highly similar, implying that consumer taxa do not shift their diet when feeding in a L. conchilega aggregation. Besides, species packing and hence trophic redundancy were not affected, pointing to an unaltered stability of the food web in the presence of L. conchilega.The potential of ecosystem engineers to modify the structure and dynamics of food webs has recently been hypothesised from a conceptual point of view. Empirical data on the integration of ecosystem engineers and food webs is however largely lacking. This paper investigates the hypothesised link based on a field sampling approach of intertidal biogenic aggregations created by the ecosystem engineer  In ecological studies, the 13C/12C and 15N/14N stable isotope ratios are the most frequently used to infer primary food sources, trophic linkages and trophic position [et al. [et al. [While the structural and functional role of biogenic reefs has been well investigated, their impact on the food web structure has only been poorly considered. Following the conceptual framework of Sanders et al. , the habposition . Generalposition , which sposition . Layman  [et al.  introduc [et al.  providedL. conchilega also causes changes in the structure of the food web. Moreover, to our knowledge, this is the first study that investigates the effect of an ecosystem engineer on the food web structure by using Bayesian Layman metrics. More specifically, we tested the hypotheses that (1) the ecosystem engineering activity of L. conchilega on its environment does not affect the isotope signatures of the primary food sources, and that (2) the altered abundance and biomass of primary producers and macrofauna in the presence of L. conchilega affects the structure, stability and isotopic niche of the consumer assemblage of a soft-bottom intertidal food web. Rather than focussing on one single ecosystem component, the entire bentho-pelagic consumer assemblage associated with the intertidal L. conchilega aggregation [This study uses stable isotopes to investigate whether changes in the species composition in the presence of regation , was takL. conchilega aggregations.Two soft-bottom intertidal areas located along the French side of the English Channel were sampled for primary food sources and consumer species. The bay of the Mont Saint-Michel (BMSM) is a large-scale intertidal sand flat located in the Normand-Breton Gulf ; while Boulogne-sur-Mer (further referred to as Boulogne) is a small-scale beach along the northern part of the English Channel . The locations were selected based on the presence of well-established intertidal L. conchilega aggregation and a (control) area in the absence of any bioengineering species. The bathymetric level between the L. conchilega aggregations and their respective control areas was similar and the sampling areas were at least 300 m apart from each other. To include temporal variability in isotopic values, sampling took place in spring 2012 (from 7th till 13th of March in the BMSM and from 22nd till 25th of March in Boulogne) and was repeated in autumn 2012 (from 17th till 21st of September in BMSM and from 15th till 18th of October in Boulogne). Sampling was conducted in cooperation with the National Museum of National History  and permitted by the ‘Affaire Maritimes’.At each location, the main primary food sources in the area (SPOM and MPB) and consumer species were sampled within a μm), the supernatant was filtered onto precombusted (450°C for 2h) and pre-weighed Whatman GF/F filters (25 mm), temporarily stored frozen at -20°C and subsequently at -80°C until processing. Fresh microphytobenthos  was collected by transferring the upper cm of the sediment to plastic boxes, covering the sediment with 100 x 150 mm Whatman lens cleaning tissue and cover slides, and putting the sediment under controlled light conditions enabling diatom migration. After about 2 days, diatoms were scraped off the cover slides, transferred to flacons with milli-Q water and centrifuged at 3000 rpm for 3 min. The diatom pellets were transferred to Sn capsules , dried at 60°C and subsequently pinch closed and stored in Multi-well Microtitre plates under dry atmospheric conditions awaiting further analysis.Sediment particulate organic matter (SPOM) was collected by sampling the upper cm of the sediment during low tide. Upon return at the lab, artificial seawater was added to the sediment and following sonication and sieving , sampling area (L. conchilega aggregation vs. control area) and period (spring vs. autumn), 3 macrobenthic cores, 1 hyperbenthic catch and 1 epibenthic catch were collected.Macrobenthic invertebrates were collected with an inox corer , sieved through a 1 mm circular mesh size and stored in a bucket with seawater. Upon return in the lab, animals were sorted, identified to the lowest possible taxonomic level, starved in seawater (24h) to allow evacuation of their gut contents and stored at -20°C before further treatment. In order to study the epi- and hyperbenthic communities, the lower water column (up to 40 cm) covering the sampling areas was sampled during daytime ebbing tide. Fish, shrimp and other epibenthic organisms were sampled with a beam trawl  equipped with a tickler-chain in the ground rope. Similarly, smaller animals living in the water layer close to the seabed  were co13C and 15N stable isotope analysis. Frozen filters were thawed, dried overnight at 60°C and subsequently acidified by exposing them to HCl fumes (37%) in a dessicator to remove inorganic carbonates [Crangon crangon, only muscular tissue  was prepared for analysis. Entire specimens and the dissected tissue samples were rinsed thoroughly with milli-Q water to avoid contamination and subsequently dried overnight at 60°C. Dried samples were grinded with a pestle and mortar, homogenized, weighted and encapsulated. The selection of the capsule depends on the need for acidification to remove carbonate traces [2 was observed [GF/F filters of SPOM and all collected consumer species were prepared for rbonates . Filterse traces , which wobserved . Followiδ notation as units of parts per thousand (‰) relative to the international reference standards:X is 13C or 15N and R is the corresponding ratio of 13C/12C or 15N/14N. Reference standards used were Vienna-Pee Dee Belemnite limestone (V-PDB) for carbon and atmospheric N2 for nitrogen. At least three replicates of SPOM and MPBdiatom were analysed for each combination of location, sampling area and period. In case of consumers, we strived to analyse at least three replicates per species, but for several taxa less replicates were available. In order to avoid pseudoreplication, every replicate equalled one single individual.In total, 399 samples  were analysed at the UC Davis Stable Isotope Facility  using a PDZ Europa ANCA-GSL elemental analyser, interfaced to a PDZ Europa 20–20 isotope ratio mass spectrometer . Stable isotope ratios are reported in the standard This study incorporated two different sampling locations (BMSM and Boulogne) along the French coastal area. Analysis of the stable isotope data was performed for each of the locations separately since they are characterized by different environmental settings.13C and δ15N isotope values of the primary food sources (SPOM and MPBdiatom) between levels of the fixed factors sampling area (L. conchilega aggregation versus control) and period (spring versus autumn) were tested by a two-way ANOVA . Significant interaction effects (p < 0.05) were further investigated by means of a TukeyHSD test. Prior to ANOVA, the assumptions of normality and homogeneity of variances were tested on untransformed data with Shapiro-Wilk tests and Levene tests respectively.Differences in the δL. conchilega aggregations and control areas were compared by plotting them in δ13C and δ15N biplots. Separate biplots were created for the locations and within biplots a distinction was made between periods. Taxa were assumed to have resembling δ13C and δ15N values in the L. conchilega and control areas if they fell within the 95% confidence interval (CI) encompassing the 1:1 correlation between L. conchilega aggregation and control isotope values.The carbon and nitrogen isotope composition of consumer taxa co-occurring in both the 13C) and trophic level (δ15N) was achieved by means of agglomerative hierarchical cluster analyses with group-average linking [15N and δ13C isotope values of individual consumers. The clusters were examined for significant differences by similarity profile (SIMPROF) permutation tests [e.g. http://www.marinespecies.org/) consumers were classified into 8 functional groups: fish, predators, omnivore/predator/scavengers, omnivores, deposit feeders/facultative suspension feeders, suspension feeders, deposit feeders and herbivores.To provide a detailed description of the entire food web structure, we used a classical approach and a more integrative approach. The classical approach includes trophic level determination and the clustering of taxa in trophic groups; the integrative approach consists of the estimation of community-wide metrics based on Bayesian statistics. Classification of consumers in groups of individuals with similar food uptake  was selected as the trophic baseline for the food webs of the BMSM, while the amphipod Gammarus sp.  for Boulogne. In addition, the use of an appropriate trophic enrichment factor (Δ15N) is important because consumers are typically enriched in their C and, mainly, N ratios relative to their prey. The generally accepted Δ15N of 3.4 ‰ was used because it was proven to be a robust and widely applicable assumption when applied to entire food webs [A useful measure for the (dis)similarity of food web structure across different systems is the trophic position (TP) of consumers in a food web, which can be estimated based on the δ5N ratio , 33:TPc crucial . As a ba crucial . The isoood webs .et al. [et al. [13C–δ 15N biplot space and measured by the first four metrics: δ15N range , δ13C range , total area of the convex hull encompassing the data  and mean distance to centroid . The extent of trophic redundancy (the relative position of taxa to each other within niche space) is measured by metrics five and six: mean nearest neighbour distance (MNND) and standard deviation of the nearest neighbour distance (SDNND). These metrics were calculated based on standard ellipses [B) was used. Standard ellipses are not sensitive to sample size because they generally contain about 40% of the data. Nevertheless, for small sample sizes (n < 30) the tendency towards underestimating the SEA remains. Therefore, a small sample-size corrected standard ellipse (SEAc), insensitive to sample size [The structure and niche widths of the food webs were investigated by calculating 6 descriptive community-wide metrics based on stable isotope data. The metrics were originally proposed by Layman et al.  and refo [et al. . Trophicellipses  and Bayeellipses . Howeverple size , was calAll univariate analyses were run in R (Version 3.1.2). The calculation of the Bayesian Layman’s metrics and standard ellipse areas for the different communities was done using SIBER .diatom (ranging from -14.08 ± 4.07‰ to -6.84 ± 0.63‰) showed a more enriched δ13C value than SPOM (ranging from -22.66 ± 0.66‰ to -20.79 ± 0.39‰) for all sampling areas and periods  to 7.21 ± 1.60‰ (MPBdiatom in the control area during autumn)  . No differaction . Howeverrol area .diatom (ranging from -19.58 ± 1.18‰ to -12.67 ± 3.97‰) showed a more enriched δ13C value than SPOM (ranging from -24.60 ± 0.70‰ to -21.21 ± 0.10‰) for all sampling areas and periods  to 4.66 ± 0.77‰  in the control area; while for Boulogne 158 organisms (42 taxa) were analysed, of which 82 organisms (34 taxa) were collected in the L. conchilega aggregation and 76 organisms (30 taxa) in the control area. The majority of organisms in the BMSM were crustaceans (52.1%) and fish (18.6%), as was the case for Boulogne (40.5% en 39.9% resp.)  and the most enriched δ13C values in the control area during autumn . Regarding δ15N, the most depleted (L. levii = 2.62‰) and most enriched (Pomatoschistus sp. = 15.93‰) values for the BMSM were found in the L. conchilega aggregation during spring, while for Boulogne the most depleted (C. crangon juvenile = 6.46‰) and most enriched (Syngnathus rostellatus = 16.68‰) values were found in the control area during autumn. In general, most taxa fell within a 1:1 correlation of isotope values between a L. conchilega aggregation and control area, i.e. their δ13C and δ15N values in the L. conchilega aggregation resembled those of the control areas  while Macoma balthica showed a depleted value in the L. conchilega aggregations. In autumn, no taxa with enriched or depleted δ15N values in the aggregation were found  inhabiting the BMSM were analysed, of which 97 organisms (36 taxa) were collected in the ganisms 4 taxa wer) Tables  and Boul) Tables , the mosol areas . Howeverre found .L. conchilega aggregation-spring and control-autumn) in the BMSM and between 4.63 (L. conchilega aggregation-autumn) and 4.97 (control-spring) in Boulogne   or at least three times as high (BMSM-spring and Boulogne-autumn) as the number of cluster in a control area. The number of functional groups between the L. conchilega aggregations and control areas was equal, but in the cases where the number of trophic groups in an aggregation were three times higher  and 5.63  than in autumn  (c) of the L. conchilega communities was larger than the those of the control communities  (B of the L. conchilega aggregation is larger than the SEAB of the control area in spring was 67.33% for the BMSM and 78% for Boulogne (c) of the L. conchilega communities were smaller than those of the control communities   . In spri12.08‰2) , Table 4 8.71‰2) , Table 4the BMSM .δ15N range (NR), the δ13C range (CR) and the standard deviation of the nearest neighbour distance (SDNND) (mean distance to centroid (CD) and the mean nearest neighbour distance (MNND) overlapped largely between L. conchilega and control communities, while slightly lower overlaps between spring and autumn communities were noted  , Table 4re noted , Table 4L. conchilega on the physical characteristics of the environment [e.g. L. conchilega does not directly affect the structure and isotopic niche of the food web of a soft-bottom intertidal ecosystem.Notwithstanding the engineering effects of ironment , 38 and ent e.g. , 20, thediatom) are the main food sources of the soft-bottom intertidal community, both in the presence and absence of L. conchilega. The tubes of L. conchilega, and animal tubes in soft bottom marine environments in general, can perturb the local flow conditions [e.g. et al. unpublished) and an increased microbial colonisation, with bacteria considered as an important food source of deposit-feeding fauna [13C values of SPOM measured in this study (-22.23 ± 1.28‰) are in the same range as δ13C values of pure phytoplankton in temperate coastal areas and estuaries [e.g. diatom on its turn is more enriched in 13C (-13.14 ± 4.39‰) compared to SPOM, which is in line with the general trend that benthic algae in coastal environments have higher δ13C values compared to phytoplankton [diatom and predominantly water column-derived suspended POM. Stable isotope values of the primary food sources were shown to be largely similar between the L. conchilega aggregations and control areas, implying an unaltered diversity of the most dominant primary resources in the presence of the tubeworm. Hence, the ecosystem engineering activity of L. conchilega does not directly modify the base of the food web. Isotope values differ rather seasonally: the δ13C value of MPBdiatom in the BMSM is higher in spring than in autumn; however the opposite is the case for SPOM. Since changes in the amount of benthic diatoms among seasons are small [in situ primary production by benthic diatoms. This finding confirms the important trophic contribution of near shore phytoplankton to sandy beach macrofauna [Primary food sources largely determine the food web structure by fuelling higher trophic levels in the system. Bulk organic matter (SPOM) and benthic diatoms  that the isotopic niche width of the L. conchilega communities is larger than the bare sand communities in spring, in combination with the deviating isotope values of some consumer taxa in the L. conchilega aggregations implies that some consumers do shift their diet preference depending on the sampling area and the period. A diet shift is for instance the case for the brown shrimp C. crangon, which is one of the most abundant species in a L. conchilega aggregation [L. conchilega aggregation in the BMSM during spring, C. crangon has a depleted δ13C value and an enriched δ15N value compared to a bare sand plot. Deviating isotope values of taxa in the L. conchilega aggregation can be the result of the uptake of a specific food source or of the different community composition in the aggregations. Firstly, the specific food uptake can be the case if a consumer feeds on a more δ13C enriched or depleted prey source which is merely available or more readily accessible in a L. conchilega aggregation due to the aggregation’s specific habitat characteristics . A prer setups, ). More mregation , 49, theL. conchilega aggregations, a more in-depth view reveals that there might be an indirect engineering effect of L. conchilega on a minor fraction of the consumer taxa owing to a specific food uptake and/or the different community composition in the aggregations.While most taxa do not show a diet shift in the L. conchilega aggregation vs. control), but differences among periods (spring vs. autumn) were shown to be slightly larger, indicating that the isotopic niche width of the consumer communities among periods is less alike than the isotopic niche width of the communities among sampling areas. A shift in the diet of the consumer taxa from spring to autumn and vice versa most reasonably explains the observed differences [L. conchilega. However, based on the increased sediment stability in the presence of L. conchilega [in situ primary production from the sediment. Comparison of trophic redundancy and overall species packing among periods reveals that the autumn communities showed an increase [by the lower values of MNND and to some extent by the lower values of CD; Isotopic niche width was not different among sampling areas  and trophic level (δ15N) for different combinations of sampling area and period in the BMSM, based on agglomerative hierarchical cluster analyses and similarity profile (SIMPROF) permutation tests. Cluster names match the clusters defined in the δ13C — δ15N biplot of the BMSM  are displayed, as well as the taxonomic composition and the number of replicates per taxon (n).Classification of consumer taxa with similar food uptake (δthe BMSM . For eac(DOCX)Click here for additional data file.S2 Appendix13C) and trophic level (δ15N) for different combinations of sampling area and period in Boulogne-sur-Mer, based on agglomerative hierarchical cluster analyses and similarity profile (SIMPROF) permutation tests. Cluster names match the clusters defined in the δ13C — δ15N biplot of Boulogne  are displayed, as well as the taxonomic composition and the number of replicates per taxon (n).Classification of consumer taxa with similar food uptake (δBoulogne . For eac(DOCX)Click here for additional data file.


In both male and female, gametogenesis is regulated by hypothalamus–pituitary–gonadal axis (HPG) that corresponds to the hormonal axis, gonadotropin-releasing hormone (GnRH)–gonadotropins–steroids. Indeed, the main target of GnRH is the gonadotrope cells, located in the adenohypophysis. These, in turn, release two gonadotropin hormones, the follicle stimulating hormone (FSH) and the luteinizing hormone (LH), that through the main circulation reach gonads to regulate gametogenesis via the synthesis of steroid hormones. It is now accepted that further non-steroid factors support germ cell progression via intragonadal action .The first evidence of relationships between pituitary and gonad came out in 1905 from a study on castrated animals, which showed hypertrophy of the pituitary gland . Later iIn the testis, it has been demonstrated that a network of intragonadal endocrine, paracrine, and autocrine factors converge in a complex stage-specific multi-factorial control of spermatogenesis . Indeed,The aim of this Research Topic is to give a comparative track on HPG axis activity for the control of spermatogenesis and quality sperm production. Through synergy between the respective specializations of all the authors, this Research Topic reviews the emerging knowledge about neuroendocrine and local mediators controlling progression and maturation of germ cells in male vertebrates.9-THC), the active principle of marijuana plant, Cannabis sativa. The middle part of this Research Topic comprises a set of four review articles dedicated to the control of fetal and postnatal development of both Leydig and germ cells and to the intragonadal networks controlling the progression of the spermatogenesis (in vitro to get a successful spermatogenesis (The Research Topic firstly reports the description of a primitive HPG in hagfish, one of the only two extant members of the class of agnathans – the most primitive vertebrates known, living or extinct – providing evidence that there are neuroendocrine–pituitary hormones that share common structure and functional features compared to later evolved vertebrates . A complogenesis –20; two ogenesis  and the ogenesis . Lastly,ogenesis , GnRH, kogenesis , and endogenesis  has beenThe last part of this Research Topic is focused on disease models such as Kallmann Syndrome , blindneWe hope that this contribution published in Frontiers in Endocrinology may represent a comprehensive guide in the plethora of data concerning the control of male reproductive activity and that readers might find new insights for the building of general models.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Livestock conservation is changing rapidly in light of policy developments, climate change, and diversifying market demands. The last decade has seen a step change in technology and analytical approaches available to define and manage Farm Animal Genetic Resources (FAnGR). However, these rapid changes pose challenges for FAnGR management in terms of technological continuity, analytical capacity, and integrative methodologies. Indeed, high-throughput single-nucleotide polymorphism genotyping is available for all major farm animal species and beyond the technological challenge to deal with these large molecular datasets, their integration with geo-environmental and socio-economic information is key to making sense of the data in practical conditions.http://www.esf.org) Research Networking Programme “Advances in Farm Animal Genomic Resources” (Genomic-Resources) proposed an action dedicated to the education of young scientists in cutting edge approaches to the characterization, analysis, evaluation, management, and conservation of FAnGR. The RNP funded three summer schools , three workshops , two conferences , and 26 exchange grants. These actions directly connected a community of 350 researchers to develop activities with the goal to meet two major challenges: (i) training in the use of novel methods able to manage and analyse high-throughput molecular data, and (ii) promoting collaboration between the animal science and social science communities to more efficiently manage FAnGR.In this context, a 4-year (2010–2014) European Science Foundation . The 31 articles can be broadly attributed to six different topic areas.In addition to the activities described above, Genomic-Resources has, in this issue of The first topic area contains general papers dealing with the identification of questions of highest priority for FAnGR research during the coming decade  and solicited contributions to this special issue to illustrate the inputs of these disciplines. Key contributions within this scope constitute part of section 3. Ahmadi et al. , comparing the genomes of European commercial pigs to their wild ancestors. The section closes with a description of SNeP, a tool to estimate changes in effective population size using genome-wide SNP data which can improve our understanding of population demography in the recent past  breeding programs. Gutiérrez-Gil et al.  compile This research topic has provided a valuable set of papers taking stock of the current advances in farm animal genomic resources worldwide. However, it inevitably lacks contributions in some areas, such as incorporation of Geographic Information Systems (GIS) to integrate complementary data on population genetics, animal husbandry practices, socio-economic and environmental characteristics (Joost et al., SJ initiated the research topic and chaired the corresponding European Science Foundation project, SJ and MB wrote and revised the manuscript, members of the Genomic-Resources consortium managed the ESF project and participated in the editorial process of this research topic.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The pooled six-, 12-, and 18-month overall survival were 69% , 36% , and 15% , respectively. Amrubicin seemed much more beneficial for Japanese patients. However, compared to the efficacy of topotecan presented in a previous meta-analysis, amrubicin may be a better treatment option than topotecan for both Japanese and Euro-American. Adverse effects by amrubicin were almost exclusively observed to be hematological. Notably, grade III/IV neutropenia incidence was 70% and febrile neutropenia incidence was 12%.Currently, amrubicin is permitted for relapsed small-cell lung carcinoma (SCLC) only in Japan. The efficacy and adverse effects of amrubicin as reported by previous studies varied greatly. The inclusion criterion was a prospective study that was able to provide data for efficacy and safety by the AMR single agent regimen as second-line chemotherapy for a patient with SCLC. Binary data were meta-analyzed with the random-model generic inverse variance method. We included nine articles consisted of 803 patients. The pooled three-, six-, and nine-month progression-free survival were 63% (95% CI 57–69%, I Small-cell lung carcinoma (SCLC) represents approximately 15% of all cases of lung cancer25Another promising chemotherapy regimen for relapsed SCLC is the amrubicin (AMR) single agent regimen289101112131415Institutional review board approval and patient consent were waivered because of the review nature of this study.st, 2015. The following search formula was used for PubMed:  AND (relapsed OR refractory OR 2nd-line OR “second line” OR 3rd-line OR “third line” OR “previously treated”) AND . The inclusion criterion for a study to be included in this meta-analysis was that it should be a prospective study that was able to provide data for at least one of following outcomes by the AMR single agent regimen as second-line chemotherapy for a patient with SCLC: objective response, progression-free survival (PFS), overall survival (OS), hematological AE, non-hematological AE, and treatment-related death (TRD). Some cases in a study could be treated as third-line chemotherapy, however, if all cases were treated as third- or later-line chemotherapy, or if any case was treated as fourth- or later-line chemotherapy, the study was excluded. Eastern Cooperative Oncology Group performance status had to be two or better. A regimen should be considered to meet current standards: 30–45 mg/m2 on day 1–3. Thus, non-standard regimens used in phase I study and a non-standard weekly regimen were excluded. The report had to be written in English language as a full article. Duplicate uses of the same data were excluded.Two investigators  systematically searched eligible articles independently. This search was conducted using the PubMed, Web of Science, and Cochrane databases as of August 1For objective response analysis, we evaluated response rate (RR) and disease control rate (DCR). If neither the number nor rate of cases who satisfied objective response and disease control were directly provided, we calculated them as follows: RR was the sum of complete response and partial response, DCR was the sum of complete response, partial response, and stable diseaseThree-month, six-month, nine-month PFS, and six-month, 12-month, and 18-month OS were evaluated. If necessary, Parmar’s method was used to estimate survival rateThe rate of AEs evaluated with National Cancer Institute-common toxicity criteria grade III or more were evaluated. Hematological toxicity such as neutropenia, thrombopenia, anemia, and febrile neutropenia; non-hematological toxicity including fatigue, nausea/vomiting, and cardiotoxicity; and TRD were evaluated. AEs were analyzed based on the number of patients, not based on the number of chemotherapy courses. The rate of AEs was evaluated using a per-protocol analysis, but not using an intention-to-treat analysis. This meant that patients who was assigned to an AMR arm but did not receive AMR were excluded from AE analysis.2 statistics was interpreted as follows: I2 = 0% indicated no heterogeneity, 0% < I2 < 25% indicated the least heterogeneity, 25% ≤ I2 < 50% indicated mild heterogeneity, 50% ≤ I2 < 75% indicated moderate heterogeneity, and 75% ≤ I2 indicating strong heterogeneityWe used the random-model generic inverse variance methodWe used meta-regression to estimate objective response and survival for each of the sensitive/refractory relapse and Japanese/Euro-American settings in the second-line scenarios19Publication bias was evaluated through visual inspection of funnel plotsRandom-effect model meta-analysis was performed using Reviewing Manager ver. 5.3 2 and one used 35 mg/m2.Of 108 articles that met the preliminary criteria, we found nine eligible articles, which included five single arm studies, three RCTs that compared AMR and TOP, and an RCT that compared AMR and a re-challenge of the first-line chemotherapy , Table 1891011121314152 = 79%) and 72% , respectively.We evaluated objective responses from 803 cases belonging to nine studies . The poo2 = 53%), 28% , 10% , respectively.Seven studies consisted of 716 patients provided data for PFS . The poo2 = 83%), 36% , 15% , respectively  for neutropenia, 24%  for thrombopenia, 19%  for anemia, and 12%  for febrile neutropenia. The pooled non-hematological AE rates were 2%  for nausea/vomitting, 7%  for fatigue, and 3%  for cardiotoxicity. TRD was observed in 1% .We evaluated AEs from 803 cases belonging to nine studies . The pooNo statistical test was conducted for publication bias because number of trials did not exceed 10. Visual inspection of funnel plots revealed no publication bias for any of the meta-analyses above .et al. recently conducted a phase III RCT to compare AMR and the current standard second-line treatment, the TOP single agent regimenThe efficacy and safety of AMR as second-line chemotherapy for SCLC has been evaluated in many phase II studies since 2006 891011121891011121314In the sensitive-relapsed Euro-American scenario, AMR can bring RR of 43% and DCR of 69% . These op = 0.047)In contrast, AMR was actually related to better OS than TOP for refractory-relapsed cases. In the refractory-relapsed Euro-American scenario, RR, DCR, six-month OS, and 12-month OS by AMR were 19%, 68%, 53%, and 19%, respectively . Accordiet al. (p = 0.047)6910111314Responses and survivals by AMR for SCLC were better in Japanese scenarios. For sensitive and refractory scenarios, 12-month OS were 51% and 34%, respectively . We do nThe safety profile of AMR is generally similar to that of TOP. AEs by AMR were almost exclusively observed to be hematological. Grade III/IV neutropenia was observed 70% of cases, which resulted in febrile neutropenia incidence of 12% .Most guidelines recommend prophylactic granulocyte-colony stimulating factor for febrile neutropenia high-risk patients when administrating a chemotherapy regimen that can improve OS in compensation for an intermediate, 10–20%, risk for febrile neutropenia2526The current study has a few limitations. First, the number of included studies was not so large. However, we successfully derived pooled outcomes adjusting baseline patient characteristics using meta-regression. Second, we could find only three trials that directly compared AMR and TOP. As of 2015, AMR is only approved in Japan; however, this medication would potentially prolong the OS of refractory-relapsed Euro-American SCLC patients. Therefore, we hope there will be further trials comparing AMR and TOP in refractory-relapsed SCLC.In conclusion, we conducted a systematic review and meta-analysis to evaluate objective response, survival, and AEs of AMR when prescribed for patients with relapsed SCLC. According to our analysis, compared to TOP, AMR provides a better objective response for both types of relapse, a similar OS for sensitive-relapsed cases, better OS for refractory-relapsed cases, and a similar AE profile except for a higher risk febrile neutropenia. AMR seemed much more beneficial for Japanese patients . HoweverHow to cite this article: Horita, N. et al. Amrubicin for relapsed small-cell lung cancer: a systematic review and meta-analysis of 803 patients. Sci. Rep.6, 18999; doi: 10.1038/srep18999 (2016).


The developing visual system of many mammalian species is partially structured and organized even before the onset of vision. Spontaneous neural activity, which spreads in waves across the retina, has been suggested to play a major role in these prenatal structuring processes. Recently, it has been shown that when employing an efficient coding strategy, such as sparse coding, these retinal activity patterns lead to basis functions that resemble optimal stimuli of simple cells in primary visual cortex (V1). Here we present the results of applying a coding strategy that optimizes for temporal slowness, namely Slow Feature Analysis (SFA), to a biologically plausible model of retinal waves. Previously, SFA has been successfully applied to model parts of the visual system, most notably in reproducing a rich set of complex-cell features by training SFA with quasi-natural image sequences. In the present work, we obtain SFA units that share a number of properties with cortical complex-cells by training on simulated retinal waves. The emergence of two distinct properties of the SFA units (phase invariance and orientation tuning) is thoroughly investigated via control experiments and mathematical analysis of the input-output functions found by SFA. The results support the idea that retinal waves share relevant temporal and spatial properties with natural visual input. Hence, retinal waves seem suitable training stimuli to learn invariances and thereby shape the developing early visual system such that it is best prepared for coding input from the natural world. It is believed that our sensory systems are adapted to statistical properties of behaviorally relevant elements in our natural environments. In the case of vision, one adaptation principle that has been put forward is the so-called slowness principle. However, the visual system is partially structured even before eye opening, when no natural input is available yet. Thus, spontaneous neural activity in the developing visual system of mammals  has been suggested to contribute to shaping connections in early visual areas before the onset of vision. Here we aim to bring these two ideas together. Specifically, we apply an algorithm that implements the slowness principle to simulated retinal waves. The algorithm is set to encode the retinal wave input and thus has to extract relevant features from that input. After encoding, we are able to investigate the emerged representation and we find that the extracted features bear strong similarity to features that are encoded by neurons in the early visual system. These features are the building blocks for an object representation that is independent of the object's position in the visual field. For us humans, vision is the most dominant sense. It is thus not surprising that the brain regions involved in visual processing, for example primary visual cortex (V1), have been subject to extensive investigation Possible answers to this question parallel the debate of nature versus nurture. One position states that the connectivity patterns are stored in the genetic code while an opposing position advocates that the connectivity patterns are acquired once the organism is confronted with its natural input. In favor of the first position would be the fact that there are animals that can see right after birth innate learning approach One such combining approach states that the visual system indeed develops to optimize a certain objective function, but it does so prior to the onset of vision by learning on internally generated input. The learning objective and the input generating mechanism would have to be stored genetically while the actual wiring of the neuronal connections can be done dynamically, driven by the objective and the input statistics. This approach includes the possibility for further learning and adaption after the onset of vision in order to maximize the organism's adjustment to its environment. Thus, this retinal wavesA prime candidate for the internally generated input to the developing visual system are the so-called As for the learning objective, there are two prominent candidates that have been applied in the context of computational modeling of the visual system, namely sparsity and slowness. Sparse coding, when trained with natural images, has been shown to lead to the emergence of functions that share relevant properties with specific types of V1 cells that are called simple cells The second candidate objective is called temporal stability, or slowness Thus, in this manuscript, we investigate to what extent it is possible to explain the emergence of complex cells based solely on two computational principles: spontaneous retinal wave activity and the slowness coding objective.g(x) that extract slowly varying scalar output signals from a high-dimensional input signal x. To ensure that the extracted output signals are informative, they are required to be uncorrelated and to have unit variance.The goal of SFA is to find instantaneous input-output functions y, respectively.The optimization problem is mathematically formulated as follows. Given a multidimensional input signal jy(t), with small Δ-values indicating a slowly varying signal. The trivial solution is a set of functions that is constant for all t. Constraints (2) and (3) avoid this trivial solution and constraint (4) ensures that different functions code for different aspects of the input signal. The latter also introduces an order, y1 being the slowest signal, y2 the second slowest and so on. We refer the reader to The expression to be minimized (1) is a measure of the temporal slowness of the signal quadratic form of the original input signal,H, f, and c are determined by SFA and the constant c is subtracted to achieve zero mean.In the present application, we chose to perform SFA in the expanded input space of all polynomials up to a degree of two, which makes it possible to conveniently express the SFA solution in terms of a g(x) to input stimuli, it is instructive to consider the eigenvalue decomposition of the matrix H. This decomposition is given by the sum over the outer products of its eigenvectors vi, weighted by the corresponding eigenvalues λi:λi and the corresponding filters are given by the eigenvectors vi.In order to understand the response properties of g(x) as there are dimensions in the expanded function space and the resulting SFA functions are ordered according to their temporal slowness, as measured on the training data. In the remainder of this article, we refer to each of the input-output functions g(x) as SFA units and limit our analysis to the 50 slowest varying functions.SFA returns as many functions π, whereas the maximal possible spatial frequency was eight cycles per receptive field, due to the 16 by 16 pixel size of the receptive field. For better comparison of the response properties between individual SFA units, the output of each unit was normalized in the following manner: First of all, the response of the unit to a gray input image was subtracted. Secondly, the output was sign corrected such that the maximal positive output is larger in magnitude than the maximal negative output. Finally, the output was normalized to have unit variance over the explored range of input stimulus parameters.Since the trained SFA units are quadratic forms of the pixel intensities, there is an explicit formulation of the input-output relation. Given a fixed-norm constraint on the input images, it is therefore possible to compute the optimal excitatory and inhibitory stimuli The full exploration of the parameter space allows us to extract for each unit the set of best parameter values, i.e. the constellation of spatial frequency, orientation and phase of the sinusoid that maximally excites the SFA unit. We visualize the units' responses as functions of the sinusoid parameters. In order to visualize these three dimensional response functions, we set one or two parameters to the values preferred by the SFA unit and plot the response as a function of the remaining parameters.response modulation indexorientation selectivity index (OSI) Additionally we employ two quantitative metrics that are used in physiological experiments as well: the The F1/F0 ratio is a spectral measure of the phase-dependence of the response. It is the ratio of the first harmonic of the response to its DC component, hence the name F1/F0 ratio. Cortical cells with an F1/F0 ratio smaller than one are classified as complex cells, whereas cells with a ratio larger than one are classified as simple cells.The OSI is a spectral measure of the orientation-dependence of the response and is given byIn order to train the SFA units, we have used input image sequences that were derived from a biologically plausible model of retinal waves proposed by Godfrey and Swindale The retinal wave model assumes that spontaneous activity of retinal amacrine cells drives the wave activity. In our simulations, the retinal amacrine cells were arranged in a regular grid consisting of 128 by 128 cells. Each cell received input from other cells that were within a six cell dendritic radius. All simulations were started with random membrane potentials and threshold levels for all cells. A cell was considered active as long as its membrane potential was above threshold. The activity of each cell was represented by a binary value and the simulated retina was visualized as a correspondingly sized image, black pixels represented non-active, white pixels represented active cells.Such an arrangement leads to inhomogeneities at the borders of the simulated patch of retina, because those cells that are situated at the borders receive less input than those that are further away from the border. Godfrey and Swindale addressed this issue by means of position-dependent regulations of activation thresholds in order to achieve a uniform distribution of wave initiation points over the retina patch. They have also reported that using cyclic boundary conditions, i.e. connecting cells at opposite borders of the simulated retina, leads to the same result. In our implementation, we chose to use cyclic boundary conditions. In order to reduce noise in the input data, spatially not coherent activity was filtered out by setting those pixels to zero that did not have more than 4 active pixels among their 8 directly adjacent pixels.After an initial warm-up phase of 30 minutes simulated time, the produced retinal activity patterns were recorded for another 15 min simulated time, which resulted in 3600 binary images (4 per second). Simulating the patch of retina is costly in terms of computational time. However, due to the cyclic boundary conditions and the otherwise homogeneous structure of the simulated retina patch, we can make an ergodicity assumption here and combine the information from several spatial locations. The image sequence obtained was tiled into overlapping receptive fields of size 16 by 16 pixels. The overlap between receptive fields was 5 pixel, resulting in 289 image sequences, each being 3600 images long and having a dimension of Prior to the quadratic expansion of the input data, the dimensionality of the concatenated image sequence was reduced to 50 by applying principal component analysis (PCA). This greatly reduces the computational costs of the SFA algorithm in terms of memory requirements. We elaborate on the effects of dimensionality reduction by means of PCA in the The trained SFA units are characterized by their optimal stimuli and their tuning to sinusoidal gratings.Once the structure of a cortical cell's optimal stimulus has been estimated in neurophysiological experiments, it allows inferences about the preferred orientation, frequency, and (in case of simple cells) the preferred phase of the cell. Simple cell optimal stimuli can be mapped by computing the spike-triggered average of random dot input stimuli and are well described by 2D Gabor functions Further response properties of the SFA units are visualized by showing their responses to sinusoidal gratings. When V1 cells are probed with sinusoidal gratings in neurophysiological experiments, the used gratings are usually parameterized along three dimensions: orientation, spatial frequency, and phase. Here we adopt the same convention and parameterize the input gratings accordingly.orientation/phase-dependent response. The spatial frequency of the gratings was set to the value that maximizes the response of the respective SFA unit. Most of the orientation/phase-dependent responses exhibit horizontal stripe patterns, indicating that the response varies stronger along the orientation axis compared to the phase axis.This rather qualitative observation was quantified using the response modulation index (F1/F0 ratio) and the orientation selectivity index (OSI). The majority of SFA units have a response modulation index smaller than one. Hence, they would be classified as complex cells in a physiological experiment. The distribution of orientation selectivity values shows a wide spread over the possible range of values, which is consistent with experimental findings The orthogonal and non-orthogonal suppression is also visible in typical orientation tuning polar plots, depicted in Most of the SFA units show a large invariance in their response with respect to phase (or position) of an input grating. Here we address the question how the SFA units achieve the phase invariance.quadrature filter pair (QFP). Recall that the SFA units are quadratic forms of the pixel intensities, in which the contribution of the quadratic and the linear term can be separately investigated , i.e. small phase invariance. H for the selected SFA units, plotted as images, as well as corresponding amplitude spectra. The last column in the figure shows the linear component.In the first column of For the most phase invariant unit (top row in the figure), the largest-magnitude eigenvectors seem to come in pairs of two. Additionally, the corresponding eigenvectors are Gabor filters with the same orientation and spatial frequency, but with a 90° phase shift. Thus, they form a quadrature filter pair. The squaring of their respective outputs and the weighting with approximately the same factor  leads to a phase invariant response when stimulated with a grating of preferred orientation and spatial frequency, as was the case here. The preferred orientation of this SFA unit is that of the eigenvector corresponding to the largest eigenvalue. Interestingly, if the orientation of the test grating deviates somewhat, the response of that unit is still large, because of the contribution of the third and fourth eigenvectors. Their preferred orientation is similar (yet wider) compared to the first and second eigenvector. This leads to a broad orientation tuning of this particular SFA unit.H that resemble Gabor filters that come pairwise and in 90° phase shift. However, the spectrum of eigenvalues does not show as many pairwise occurring eigenvalues as was the case in the most phase invariant unit. Thus, the squared output of the corresponding filters in the quadrature filter pairs are not as equally weighted and thereby leads to oscillations in the output of The second-most phase invariant SFA unit (middle row) also shows eigenvectors of The eigenvectors of the third unit shown (bottom row) do not resemble phase shifted pairs of Gabor filters. The lack of appropriate filters and corresponding pairwise eigenvalues leads to strong oscillations in the phase dependent response curve, i.e. very little phase invariance. Such a cell may, however, be very invariant to other features , which were not varied in this particular test.To summarize, we have found that the phase invariance of the SFA units is explained by two features, namely (1) the emergence of pairs of Gabor filters with equal orientation and spatial frequency but 90° phase shift, and (2) the emergence of correspondingly pairwise eigenvalues, which together constitute Gabor quadrature filter pairs. This property is observed in about half of the SFA units obtained.The histogram of OSI values in Recall that the OSI measures the ratio between the second harmonic of the orientation tuning curve (F2) and the sum of F2 and the average response across all orientations (F0). Thus two scenarios can lead to large a OSI: Firstly, the OSI becomes large if there are two peaks in the orientation tuning that are separated by 180° and the response to other directions is comparably low. Secondly, the OSI becomes large if the average response (F0) is close to zero. In fact, the OSI is maximal if F0 equals zero, independent of how orientation selective the analysed SFA unit actually is.In the SFA units obtained, both effects work together and thereby yield large OSI values. The presence of rotation in the training data is predicted to lead to harmonic oscillations in the orientation tuning curve Interestingly, there is another mechanism that could lead to smooth orientation tuning curves of SFA units, namely the combination of (i) finite receptive field size and (ii) input dynamics that are dominated by translation. In the case where the training input contains only translation and no rotation at all, the theory behind SFA predicts a rather erratic orientation tuning In order to answer this question we have conducted additional control experiments. Specifically, we conducted simulations in which SFA units were trained with pink noise image sequences that contained either only translation, only rotation, or a combination of translation and rotation applied to the images. A similar simulation protocol was used in Additionally, we analytically derived the OSI for a complex cell model that consists of a number of Gabor patch quadrature filter pairs (Gabor-QFP model), see From this we conclude that the smoothly varying orientation tuning curves we obtained from retinal waves are not an artifact of finite receptive field size. Instead, the orientation tuning is a direct result of the SFA units adapting to rotational components in the input data.In this article, we present the results of applying slow feature analysis to image sequences derived from a model of retinal waves. The resulting SFA units share a number of properties with complex cells, which are found in adult mammalian primary visual cortex. The defining feature of cortical complex cells is that they respond well to sinusoidal gratings and show little variation in their response when the phase of the grating changes. This most important feature can be reproduced with the SFA model, i.e. the response of the SFA units is largely invariant with respect to the phase of a sinusoidal input grating. Secondly, the optimal stimuli of a large portion of the SFA units show structure similar to that found in experimentally observed receptive fields. Many optimal stimuli resemble Gabor patches. Thirdly, similar to cortical simple and complex cells, the SFA units respond stronger to some orientations of the input grating than to others, i.e. they exhibit an orientation tuning. However, some specifics of their orientation tuning are not in accordance with physiological observations (see Result section and below for further discussion of this issue). Finally, the SFA units learned exhibit some degree of frequency tuning, which is also found to be the case in cortical simple and complex cells.Previously, models based on temporal slowness have been applied to natural image sequences and have reproduced properties that are reminiscent of cortical complex cells Albert et al. have applied sparse coding The retinal wave image sequences used for training the SFA units were derived from a model of retinal waves proposed by Godfrey and Swindale 9, while for 50 input dimensions the number of entries to estimate is in the order of 106.Recall that we have applied PCA dimensionality reduction to the simulated retinal wave image sequences before using them as input to SFA. Dimensionality reduction was a necessary step for reasons of computational efficiency. SFA acts on second moment matrices that have to be estimated on the quadratically expanded input data. For 256 input dimensions, the number of entries of these matrices is in the order of 10Applying PCA for dimensionality reduction is essentially equivalent to spatially low-pass filtering the input, due to the characteristic fall-off of power in the Fourier spectrum of natural images, as well as the spectrum of simulated retinal waves . Yet the dimensionality reduction allows to use larger receptive fields compared to using unprocessed input. Larger receptive fields in turn allow for an easier analysis, especially with respect to the interpretation of optimal stimuli. Berkes and Wiskott have thoroughly discussed the effects of PCA dimensionality reduction prior to SFA in However, using PCA processed (or low-pass filtered) image patches limits the extent of frequency-specific response of any subsequent coding mechanism. Similar to Berkes and Wiskott The SFA algorithm as such is not intended to be biologically plausible in any detail. It is an abstract implementation of the slowness principle. Thus SFA may produce effects that an actual neural system would not, or it may fail to reproduce certain aspects of actual neural system responses. However, SFA is a self-organizing learning algorithm which implements the slowness principle and thus it is expected to reproduce properties of neural systems that are directly related to the coding objective. In the present application, that particular property is phase invariance. The phase invariance of cortical complex cells can be well reproduced, which renders SFA a suitable model for the emergence of such. Furthermore, the SFA units obtained show direction- and frequency preference as well as secondary response lobes. In addition to the properties reproduced here, the modeling study of Berkes and Wiskott Note that the standard complex cell model that consists of a single quadrature filter pair only is unable to produce response properties such as secondary response lobes or end- and side-inhibition. More than one pair is necessary to achieve these properties, which indicates that while the standard single quadrature pair model is sufficient to account for phase invariance and orientation selectivity, it is too simple a model to account for the properties previously mentioned. The SFA model, on the other hand, is complex enough to enable the emergence of multiple quadrature filter pairs.The observed emergence of multiple quadrature filter pairs is in line with the analytical derivation of complex cell properties from the slowness principle presented in The orientation selectivity exhibited by the SFA units obtained in the present study is well above experimental findings when measured with the orientation selectivity index OSI To the best of our knowledge, there are no studies that describe the properties of complex cells shortly after or shorty before birth. The experimental findings cited in this work are primarily derived from simple cells. However, the results of our work predict that complex cells could already be present at the time of birth or shortly after, possibly in some preliminary form. It seems likely that this is the case at least in some animal species, such as horse or giraffe, because their offspring is born at a very advanced developmental stage. The freshly born foals or calves are able to stand and follow their mother within the first hour after birth. It seems likely that these animals, even at this young age, can recognize and differentiate objects (e.g. other members of their species) independently of their position in the visual field, possibly using also other sensory cues such as olfactory or auditory signals. Complex cells are a prime candidate for the basis of position invariant object recognition Given that our first prediction can be experimentally verified, i.e. if it becomes possible to record from complex cells in animal species before or shortly after the onset vision, then a second, more principled, prediction concerning the emergence of simple and complex cells can be derived. The performed control experiments demonstrate the dependence of the SFA units on the transformations that are present in the training stimuli. If one could disrupt the temporal structure of retinal waves while leaving the spatial properties intact, then, according to the theory behind SFA, the development of complex cells should be impaired. The development of simple cells, on the other hand, should not be hindered, because the theory of sparse coding is based on spatial statistics only, making no reference to the temporal properties of the training data. Currently the necessary technology may not be available to actually perform such an experiment. However, it is already possible to pharmacologically block natural retinal waves The results presented in this study can only give a rather simplified account of the processes that come together in structuring the neural circuitry of sensory systems. Specifically, our study addresses only a particular part of the development of the early visual system, namely complex cells. Already the clear-cut distinction between simple and complex cells is most likely too sharp, as it seems more likely to be a continuum between purely simple and purely complex cells With the present study we aim at showing how far one can get from two simple computational principles (endogenous activity and the slowness coding objective). Our results indicate that a number of interesting properties can emerge from these simple ingredients already, yet they cannot account for the true complexity of our intricate neural system.In conclusion we find that our simulation results support the hypothesis that the slowness objective, here manifested by SFA, is compatible with an innate learning mechanism that learns on endogenous activity in the same manner as on actual visual input. A large portion of the SFA units obtained from training with retinal wave image sequences share relevant properties with cortical complex-cells. Thereby we provide a theoretical account for the emergence of complex-cells prior to eye opening that allows for further refinement by exposure to natural visual input.Text S1In this Supporting Information we provide the analytical derivation of a formula for the orientation selectivity index (OSI) of a linear combination of Gabor wavelet quadrature filter pairs (Gabor-QFPs). The derived formula is the basis for sampling OSI values from the Gabor-QFP model and generating the OSI distribution shown in (PDF)Click here for additional data file.


Drosophila dopamine transporter (dDAT) gene show enhanced dopamine signaling, and short sleep and memory impairment phenotypes. However, understanding the mechanism by which dopamine signaling causes these phenotypes requires an understanding of the dynamics of dopamine release. Here we report the effects of dDAT expression on behavioral traits. We show that dDAT expression in a subset of dopaminergic neurons is sufficient for normal sleep. dDAT expression in other cell types such as Kenyon cells and glial cells can also rescue the short sleep phenotype of dDAT mutants. dDAT mutants also show a down-regulation of the D1-like dopamine receptor dDA1, and this phenotype is rescued when dDAT is expressed in the same cell types in which it rescues sleep. On the other hand, dDAT overexpression in mushroom bodies, which are the target of memory forming dopamine neurons, abolishes olfactory aversive memory. Our data demonstrate that expression of extrasynaptic dopamine transporters can rescue some aspects of dopamine signaling in dopamine transporter mutants. These results provide novel insights into regulatory systems that modulate dopamine signaling.Dopamine mediates diverse functions such as motivation, reward, attention, learning/memory and sleep/arousal. Recent studies using model organisms including the fruit fly, have elucidated various physiological functions of dopamine, and identified specific neural circuits for these functions. Flies with mutations in the  DAT) mediates reuptake of dopamine from the synaptic cleft, and has a key role in limiting dopamine signaling. DAT is inhibited by therapeutic agents such as Ritalin (methylphenidate) or psychostimulant drugs of abuse such as amphetamine and cocaine, demonstrating its importance in regulating behaviors. Besides directly regulating dopamine signaling, DAT is thought to participate in gating the spillover of dopamine , sleeps less than wild-type flies due to an increase in postsynaptic dopamine signaling , abolished olfactory memory formation. These results shed light on the physiological function of dDAT in dopamine signaling and in regulating extrasynaptic dopamine spillover in vivo.In this report, we describe effects of altered fmn (fumin) mutant was described previously  line for DAT (Transformant ID: 106961), UAS-Dicer-2 flies (Stock numbers: 60008) and the w1118 (60000) which is the genetic background of the RNAi line were obtained from the Vienna Drosophila RNAi Center (VDRC), Vienna, Austria . Based on previous reports  containing 1% agar and 5% sucrose at 24.5°C. Male flies were used for sleep analysis. They were entrained for at least 3 d to LD conditions before being transferred to constant dark (DD) conditions. Locomotor activity was monitored by recording infrared beam crossings by individual flies in 1 min bins using the dDAT RNAi were examined by qPCR. Total RNA was extracted from 10 heads by using RNeasy Micro Kit (Qiagen) according to the manufacturer’s instructions. cDNA was synthesized from the total RNA using oligo (dT)20 primer and ReverTra Ace reverse transcriptase (Toyobo). The cDNA was used for qPCR using THUNDERBIRD SYBR qPCR Mix (Toyobo). The primers were 5′-AACAATAGCA TCAGCGACGA-3′ and 5′- CAGGTTATGG CACCCTTACG-3′ for dDAT, 5′- TGGTACGACA ACGAGTTTGG-3′ and 5′- TTTCAGGCCG TTTCTGAAGT-3′ for GAPDH2. Before qPCR, all the primer sets were confirmed to yield one major band corresponding to each targeted mRNA on the gel when used in conventional RT-PCR. Each expression level was first normalized to GAPDH2. Then, the values were normalized to the average of independent control sample.The efficiency of Brains of adult flies were dissected in cold PBS and fixed in 4% paraformaldehyde in PBS for 20 min at room temperature. After three 20 min washes in 0.3% Triton X-100 in PBS, samples were blocked and penetrated in 5% normal sheep serum (NSS) for 30 min at room temperature. Samples were then incubated in a primary antibody solution in 5% NSS at 4°C overnight. The following primary antibodies were used in this study: chicken anti-GFP , rabbit anti-TH (1:25 Pel-Freez), rat anti-DAT  and rabbit anti-dDA1 (1:1250: gift from Dr. Fred W. Wolf). After three 20 min washes in 0.3% Triton X-100 in PBS, brains were incubated at 4°C overnight in a secondary-antibody solution in 5% NSS. The following secondary antibodies were used: Alexa Fluor 568 goat anti-rat IgG , Alexa Fluor 488 goat anti-chicken IgG  and Alexa Fluor 568 goat anti-rabbit IgG . After three 20 min washes in PBS, brains were mounted using PermaFluor (Thermo Scientific) between two coverslips separated by electrical tape of ~200 μm thickness, so that the brain sample was not flattened. Immunolabeled adult brains were imaged under a Leica TCS SP2 AOBS confocal microscope using 20x magnification. Z-stack images were scanned at 2 μm section intervals with a resolution of 1024 × 1024 pixels.For quantification of dDA1 signal, a confocal slice image containing the holizontal lobes of the mushroom body was selected from a stack. The average pixel intensity values in the horizontal lobes and the adjacent control region  was measured by NIH ImageJ. The signal of the dDA1 in horizontal lobes was normalized by the control region of an identical confocal slice.Olfactory conditioning with two odors (4-methylcyclohexanol and 3-octanol) was performed as described previously  or analysis of one-way ANOVA with Tukey-Kramer HSD post hoc test (for more than two groups). Bars and error bars represent means and SEM, respectively. Data were analyzed as described in the figure legends using R 2.11.1.The significance level in each experiment was set to 5%. Comparisons were made by using either Student’s dDAT mutant fumin shows a short sleep phenotype , glial cells (dEAAT1-GAL4) and pars intercerebralis (c767-GAL4) increased dDA1 expression in the mushroom body, indicating decrease of dopamine signaling. These results are consistent with the rescued sleep phenotype in fmn by ectopic dDAT expression  as well as dopamine neurons (using TH-GAL4) promoted memory formation in fmn mutants although we could not find statistical significance. Surprisingly, however, when some GAL4 drivers which are expressed in the mushroom body (such as 30Y-GAL4 and OK107-GAL4) were used to drive dDAT, these flies did not demonstrate aversive olfactory memory . Flies with aaNAT mutation show defects in sleep homeostasis  expression in the fly brain and suggested additional factors may be needed for dSERT function . Cell-type-specific ectopic expression of transporters for neurotransmitters in Drosophila will be a favorable tool by which the mechanisms underlying precise inter-cellular signaling in space and time will be elucidated.Since synaptic and extrasynaptic glutamate uptake transporters are enriched on multiple presynaptic and postsynaptic cells , neurotransmitters such as glutamate are restricted to the synaptic cleft of the release site (Seal and Amara, A previous study explored the relative roles of the DAT on striatal extracellular dopamine during tonic vs. phasic activity of dopamine neurons in the substantia nigra (Floresco et al., The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


This conceptual framework and first empirical data provide a spring-board for directing future genomic analyses on exploiting social parasites as a route to understanding the evolution of phenotypic specialization.Contrasting phenotypes arise from similar genomes through a combination of losses, gains, co-option and modifications of inherited genomic material. Understanding the molecular basis of this phenotypic diversity is a fundamental challenge in modern evolutionary biology. Comparisons of the genes and their expression patterns underlying traits in closely related species offer an unrivaled opportunity to evaluate the extent to which genomic material is reorganized to produce novel traits. Advances in molecular methods now allow us to dissect the molecular machinery underlying phenotypic diversity in almost any organism, from single-celled entities to the most complex vertebrates. Here we discuss how comparisons of social parasites and their free-living hosts may provide unique insights into the molecular basis of phenotypic evolution. Social parasites evolve from a eusocial ancestor and are specialized to exploit the socially acquired resources of their closely-related eusocial host. Molecular comparisons of such species pairs can reveal how genomic material is re-organized in the loss of ancestral traits  and the gain of new ones . We define hypotheses on the molecular basis of phenotypes in the evolution of social parasitism and discuss their wider application in our understanding of the molecular basis of phenotypic diversity within the theoretical framework of phenotypic plasticity and shifting reaction norms. Currently there are no data available to test these hypotheses, and so we also provide some proof of concept data using the paper wasp social parasite/host system  Evolution plays with inherited traits to produce altered phenotypes which may be better adapted to fill a niche different to that of their ancestors. Ultimately, phenotypic traits arise at the level of the genes. A major outstanding question in evolutionary biology is what roles do losses, gains, co-options and modifications of genomic material play in the evolution of phenotypic diversity within and between species?  and Alpinobombus,  representing over 11 independent origins of eusociality. Their societies are defined by a division of reproductive labor in the form of queen and worker castes, overlapping of generations, and cooperative brood care. Social parasitism has evolved multiple times independently in the eusocial insects: three times in wasps  or loose (B) sense, and thus share recent genomic ancestry : Acromymex insinuator , Acromyrmex echinator (the host), and Acromyrmex octospinosus (non-host sister species); (B): Polistes sulcifer , Polistes dominula (the host) and Polistes nimphus (non-host sister species). This is important for genomic analyses of phenotypic plasticity where we are interested in understanding the molecular basis of traits other than morphology . Shared morphology between parasite and host therefore helps to controls to some extent for the machinery underlying morphological differences. Molecular analyses also help with social parasite species discovery, as the parasites may be cryptic at the morphological level, but not at the molecular level.Cryptic morphology. Although hymenopteran social parasites differ significantly to their hosts in life strategy and behavior, they are usually near indistinguishable from their hosts morphologically e.g., , but also exhibit novel traits . Whilst these are well studied at the phenotypic level, we know nothing about how such losses and gains occur at the molecular level. Comparisons of the molecular bases of closely related host and social parasite traits will provide new insights into phenotypic evolution.Photo credits: Alessandro Cini, Rita Cervo, Stefano Turillazzi and David Nash.Eusocial species evolve from solitary ancestors. Solitary phenotypes occupy a normal distribution of variation, determined by their individual threshold level of response to environmental cues Figure . Queen aHere we present some testable hypotheses for these models. These hypotheses and predictions are specific to obligate social parasites and their eusocial insect hosts, but they may also be of general relevance to furthering our understanding of the molecular basis of phenotypic diversity. The empirical approach we suggest requires a combined analysis of individual-level behavioral monitoring with subsequent quantitative analyses of the many components of the molecular phenotype , because they are both reproductive specialists. Support for this hypothesis would suggest that social parasites are simply a reduced form of the social phenotype, expressing the reproductive component, but suppressing the worker component of their ancestors , or a response to a new cue .Molecular phenotypes  are highly labile and can change responsively to environmental variation. A key question is whether different organisms use the same genes to respond to the same environmental cues. There will be strong selection for the social parasites to be able to accurately detect and respond to their host's environmental cues since they share the same intimate environment on the nest. Moreover, the social parasite must synchronize its life cycle and behavior perfectly with the host's life cycle Cervo, . The moll Figure  is compaOne important phenotype-environment response trait in both hosts and social parasites is the ability to respond to the switch from a solitary to social environment. Many eusocial insects have a solitary phase, when a single queen founds a new colony and raises her first brood alone, and then switches to a eusocial phase when her workers emerge see Box . LikewisBox 2P. sulcifer is the obligate social parasite of its close relative, the primitively eusocial wasp P. dominula  is a putative candidate gene for response to changes in the social environment. We observed up-regulation of IGF in social parasites brains when they shift from solitary to social living, whilst IGF shows no change in expression in the constant eusocial environments of the host , usurping (U) and post-usurpation (SU) phases of their life cycle (see Box Art) and Apolipophorin (Apo)  Figure . These g) Figure . During ) Figure , which iProtomognathus americanus ants  queen . Social parasites therefore may not evolve through simple “deletion” (silencing) of the worker phenotype and its associated molecular functions (West-Eberhard, Nasonia, Werren et al., Our model and predictions are preliminary, but are relevant more widely to non-hymenopteran social parasites, as social parasitism of parental care has evolved multiple times in different taxa of the animal kingdom, e.g., birds Davies, ; lycaeniThe authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.