Ephx2 knock-out (KO) mice exhibit stress resilience after chronic social defeat stress. Furthermore, the sEH inhibitors have antidepressant effects in animal models of depression. In addition, pharmacological inhibition or gene KO of sEH protected against dopaminergic neurotoxicity in the striatum after repeated administration of MPTP  in an animal model of Parkinson’s disease (PD). Protein expression of sEH in the striatum from MPTP-treated mice was higher than control mice. A number of studies using postmortem brain samples showed that the deposition of protein aggregates of α-synuclein, termed Lewy bodies, is evident in multiple brain regions of patients from PD and dementia with Lewy bodies (DLB). Moreover, the expression of the sEH protein in the striatum from patients with DLB was significantly higher compared with controls. Interestingly, there was a positive correlation between sEH expression and the ratio of phosphorylated α-synuclein to α-synuclein in the striatum. In the review, the author discusses the role of sEH in the metabolism of PUFAs in inflammation-related psychiatric and neurological disorders.Inflammation plays a key role in the pathogenesis of a number of psychiatric and neurological disorders. Soluble epoxide hydrolases (sEH), enzymes present in all living organisms, metabolize epoxy fatty acids (EpFAs) to corresponding 1,2-diols by the addition of a molecule of water. Accumulating evidence suggests that sEH in the metabolism of polyunsaturated fatty acids (PUFAs) plays a key role in inflammation. Preclinical studies demonstrated that protein expression of sEH in the prefrontal cortex, striatum, and hippocampus from mice with depression-like phenotype was higher than control mice. Furthermore, protein expression of sEH in the parietal cortex from patients with major depressive disorder was higher than controls. Interestingly,  Polyunsaturated fatty acids (PUFAs) are generally considered to be necessary for maintaining normal physiology . PUFAs aPolyunsaturated fatty acids (PUFAs) are metabolized into bioactive derivatives by the main enzymes such as cyclooxygenases (COXs), lipoxygenases (LOXs), and cytochrome P450s (CYPs) . The COXIn the review, the author would like to discuss the role of soluble epoxide hydrolase (sEH) in the CYP-mediated metabolism of PUFAs which might be involved in the pathogenesis of psychiatric and neurological disorders. Furthermore, we also refer to the clinical significance of sEH inhibitors for these disorders.The CYP system is a superfamily of enzymes, which are involved in the metabolism of exogenous and endogenous compounds. The CYP in the eicosanoid pathway was first described in 1980 and is comprised of two enzymatic pathways such as hydroxylases and epoxygenases. The CYP isoforms metabolize a number of ω-3 and ω-6 PUFAs, including ARA, EPA and DHA into bioactive lipid mediators, termed eicosanoids . The CYPEPHX2 gene codes for the sEH protein is widely expressed in a number of tissues, including the liver, lungs, kidney, heart, brain, adrenals, spleen, intestines, urinary bladder, placenta, skin, mammary gland, testis, leukocytes, vascular endothelium, and smooth muscle. Interestingly, the sEH protein is most highly expressed in the liver and kidney  . The sEHd kidney .Accumulating evidence suggests that EETs, EDPs and some other EpFAs have potent anti-inflammatory properties  which arDepression, one of the most common disorders in the world, is a major psychiatric disorder with a high rate of relapse. The World Health Organization (WHO) estimates that more than 320 million individuals of all ages suffer from depression . MultiplSeveral reports using meta-analysis demonstrated that ω-3 PUFAs could reduce depressive symptoms beyond placebo . DietaryGiven the role of inflammation in depression, it is likely that sEH might contribute to the pathophysiology of depression. A single injection of lipopolysaccharide (LPS) is known to produce depression-like phenotypes in rodents after sickness behaviors . Ren et A study using euthymic patients with a history of major depressive disorder with seasonal depression showed changes in CYP- and sEH-derived eicosanoids in patients with winter depression . The ω-6EPHX2) gene was found to harbor several common and rare risk variants for AN  is a serious eating disorder characterized by the persistent restriction of energy intake, intense fear of gating weight, and distribution in self-perceived weight or shape. The Epoxide Hydrolase 2 (s for AN . Subsequs for AN . Interescy in AN . These dcy in AN .Attention deficit hyperactivity disorder (ADHD) is one of the most common psychiatric disorders affecting children. Symptoms of ADHD include inattention, hyperactivity and impulsivity. However, biological mechanisms underlying ADHD remain unknown. A meta-analysis shows that children and youth with ADHD have elevated ratios of both blood ω-6/ω-3 PUFAs compared to controls , suggestParkinson’s disease (PD) is the second most common neurodegenerative disease after Alzheimer’s disease. Although the precise pathogenesis of PD is unknown, the pathological hallmark of PD involves the progressive loss of dopaminergic neurons in the substantia nigra (SN) . In addiω-3 PUFAs appear to be neuroprotective for several neurological disorders. It is reported that dietary intake of PUFAs is associated with lower risk of PD . MPTP 1. A diet Deposition of α-synuclein has been shown in multiple brain regions of PD and DLB patients . InteresPARKIN gene , showed an IC50 of 92 nM via fluorescent assay and a Ki of 54 nM via radioactivity-based assay on human sEH urea], the mosuman sEH . MMU is uman sEH . These fAnother topic is the systemic anti-inflammatory effects of sEH inhibition or genetic disruption . TherefoIn conclusion, considering the role of sEH in the metabolism of EpFAs , treatment of ω-3 PUFAs in combination with a sEH inhibitor could represent a novel therapeutic approach for psychiatric and neurological disorders. This approach may well bridge the currently unmet medical needs for these CNS disorders.The author confirms being the sole contributor of this work and has approved it for publication.The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Here, we use group-theoretical analysis to uncover a profound enhancement of the number of improper ferroelectric coupling schemes available to molecular perovskites. This enhancement arises because molecular substitution diversifies the range of distortions possible. Not only do our insights rationalise the emergence of polarisation in previously studied materials, but we identify the fundamental importance of molecular degrees of freedom that are straightforwardly controlled from a synthetic viewpoint. We envisage that the crystal design principles we develop here will enable targeted synthesis of a large family of new acentric functional materials.The central goal of crystal engineering is to control material function via rational design of structure. A particularly successful realisation of this paradigm is hybrid improper ferroelectricity in layered perovskite materials, where layering and cooperative octahedral tilts combine to break inversion symmetry. However, in the parent family of inorganic ABX In layered inorganic materials lattice distortions can couple to break inversion symmetry and drive improper ferroelectricity, but in perovskites, symmetry prohibits such an effect. Here, the authors use group-theoretical analysis to provide crystal engineering design principles for improper ferroelectricity in molecular perovskites. An early canonical ferroelectric was BaTiO3, with the broader family of ABX3 perovskite oxides now known to include a variety of ferroelectric systems5. The ferroelectric response of BaTiO3 originates from a second-order Jahn–Teller (SOJT) effect, in which the B-site cation Ti4+ displaces from the centre of its TiO6 coordination environment6. As this polar instability also acts as the primary order parameter, BaTiO3 is a proper ferroelectric. Despite the continuing discussion regarding the origin of polar coupling in BaTiO37, the general SOJT mechanism at play is rare and its generalisation is challenging, as it requires a d0 electronic configuration6. Hence, design approaches for new ferroelectric materials based on SOJT instabilities are limited and, moreover, the mechanism is difficult to couple with spin-active d-electron configurations, which has largely prevented its exploitation in the search for magnetoelectric multiferroics8. In fact, ferroelectricity is comparatively rare in bulk perovskite materials and BaTiO3 is much more of an exception than a rule9.Ferroelectricity, i.e., the presence of a switchable electric polarisation, is an important property with many technological applications10. A necessary condition for ferroelectricity is the presence of a polar space group, which in turn requires broken inversion symmetry. Simple inorganic perovskites contain two crystallographically distinct inversion centres  and in BaTiO3, the zone-centre polar mode breaks both of these, driving the polarity11. Alternatively, in favourable cases, a combination of two or more modes—each non-polar in their own right—may collectively lift inversion symmetry and give rise to a polar secondary order parameter12. This so-called hybrid improper ferroelectricity mechanism is attractive from a crystal engineering perspective because it lends itself to design rules via group-theoretical analysis10. In addition, the mechanism does not preclude magnetic order13. Hence, using group-theoretical methods, it is possible to enumerate the symmetry breaking caused by given distortions of an aristotype and thereby predict the propensity for the formation of acentric structures. For simple inorganic perovskites, the accessible degrees of freedom—cooperative first-order Jahn–Teller (FOJT) distortions and octahedral tilting—all preserve the inversion centre of the B-site, and in order to enable hybrid improper ferroelectricity, additional symmetry breaking in the form of A-site cation order or layering is needed15.It is in this context that the concept of hybrid improper ferroelectricity is especially appealing16. These are solid compounds with the same ABX3 stoichiometry of conventional perovskites, but where A or X (or both) are molecular ions. This populous class of materials may be categorised according to the nature of the anionic linker: topical families include the organic–halide perovskites18, metal formates20, Prussian blue analogues22, azides24, dicyanamides26, hypophosphites27, thiocyanates29, and dicyanometallates31. The enhanced structural flexibility allowed by molecular species enables additional degrees of freedom unfeasible in conventional inorganic perovskites: unconventional tilts32, columnar shifts33, and multipolar order 36—i.e., phonon modes which propagate without deforming BX6 coordination geometries38. The larger number of RUMs in molecular perovskites is conceptually related to the additional flexibility driven by reduced connectivity in Ruddlesden–Popper phases39. By contrast, the presence of multipolar degrees of freedom reflects reorientations of non-spherical molecular A-site cations40.A recent development in the broader field is the increased interest in molecular perovskite analoguesIn this paper, we show how these new types of structural degrees of freedom can combine to break inversion symmetry, hence establishing a new set of design rules for engineering acentric molecular perovskites. First, we introduce our general recipe for designing acentric materials. Second, we classify the various symmetry breaking ingredients accessible to molecular perovskites in terms of the irreducible representations of the high-symmetry perovskite aristotype. Third, we use these ingredients to construct the key ferroelectric coupling schemes. And, fourth, we illustrate how this approach can be used to rationalise the emergence of polarisation in a number of previously reported systems.A and B) of the parent P) in the hettotype (child structure). In a Landau-style expansion of the free energy about the parent structure, this produces a third-order (trilinear) term βABP. As A and B are inherently non-zero if they are unstable with respect to the parent phase then—irrespective of the sign of the coefficient β—P will also adopt a non-zero (positive or negative) value in order to stabilise the free energy. We can use the idea of invariants analysis41 to identify the permissible third-order terms in the general free energy expansion; i.e., identify for which combinations of A and B one would expect coupling to P. For the zone-boundary/zone-centre distortions that we consider here, we find that such couplings may in fact be identified by inspection, as detailed in the following paragraphs.The fundamental idea of our approach is to identify two distortions , Each distortion we consider can be described as transforming as an irreducible representation (irrep) of the parent space group mentclass2pt{minimmentclass2pt{minimmentclass2pt{minim+ and X−, or as M+ and M−, or as R+ and R− must couple to a non-centrosymmetric  distortion P, which itself transforms as Γ−. On the other hand, any two distortions to the parent phase that have either a different propagation vector and/or the same sign with respect to inversion parity cannot produce such a coupling to P. Hence, combinations such as X+ and R−, or M− and M− etc., may be immediately discounted. As a final point, we note that higher-order coupling terms such as , , and so on also produce a fourth-order coupling in P, provided that the rules for preservation of crystal momentum and parity are observed.As any term that features in the free energy expansion about the parent phase must conserve both crystal momentum and parity with respect to the inversion symmetry, it can quickly be seen that any two distortions transforming as XIn the context of molecular perovskites, the relevant collective distortions involve conventional tilts, unconventional tilts, columnar shifts, FOJT effects, and multipolar A-site order. The combinations of distortions capable of driving polarity are shown in Fig. Arguably one of the easiest distortions to control from a materials design perspective is that of A-site multipolar order. Many common organic cations are polar—their charge being localised on a single atom. Moreover, both rod-like (prolate) and disc-like (oblate) cations possess quadrupole moments. This includes common cations such as methylammonium mentclass2pt{minimBut to what extent are these predictions borne out in practice? There are in fact a number of hybrid ferroelectric molecular perovskites already reported in the literature. In Table 4]Cd(HCOO)3, first synthesised several decades ago42 and revisited more recently for its dielectric properties43. The system crystallises in the polar space group Pna21, with the formate anion adopting a mixed syn-anti binding mode. This binding geometry is less prevalent than the anti-anti binding mode typically observed for formate-bridged perovskites47, and is related to the low tolerance factor47. The structure contains a considerable planar shift alternating along the c direction which is described by the irrep Cmc21, which in combination with the conventional in-phase tilting Pna21. Despite the fact that three order parameters are needed to fully account for the observed space group, the shift Our first experimental example is that of the formate perovskite [NH3 (where Gua = Pna21 space group symmetry. In this case the coupling is between a cooperative Jahn–Teller distortion and multipolar A-site order. It is the only polar member of the family [Gua]M(HCOO)3 48, and also the only Jahn–Teller-active member of the same family, which immediately identifies the importance of a collective Jahn–Teller distortion 350. Quantum mechanical calculations estimate the maximum polarisation in this system to be 0.22 μC/cm2 50, which demonstrates that the achievable polarisations in these molecular perovskites is likely comparable to that of the Rochelle salts, for example51.Our second example is that of [Gua]Cu(HCOO)3 with pre-existing structural distortions  such that superposition of JT order would be expected to give a polar state. In addition to the guanidinium transition-metal formates discussed above, we identify two additional molecular perovskites that satisfy this criterion for 2PO2)3 3 , which is isostructural and hence described in terms of the same distortion modes52. In both cases, substitution of Mn for Cu might reasonably be expected to drive hybrid improper ferroelectricity—a prediction that could be tested experimentally.Drawing on this concept of inversion symmetry breaking via coupling to collective JT order, we flag the possibility of identifying ‘propolar’ molecular frameworks: i.e., systems such as [Gua]M(HCOO)Pnma53, it is not yet clear what crystal symmetry—if any—is especially prominent amongst molecular perovskites16. Hence, we illustrate this strategy with the Pnma perovskite structure, mindful of the possibility of extrapolating to other structures as our collective understanding develops. The key distortions in the Pnma structure are two tilts—one in-phase Pnma perovskite, will give rise to hybrid improper ferroelectricity, provided that the dipole moment orders at the M-point  or the quadrupole moment orders at the X- or R-point. To maximise the likelihood of such a situation, one should choose cations with both quadrupolar and dipolar moments, of which there are many examples; e.g., hydrazinium, imidazolium, and methylammonium. Steric considerations will stabilise orientational order of larger cations to lower temperatures, and design rules for controlling the type of quadrupolar order are now starting to emerge35. Hence, judicious choice of a molecular cation with the correct type of multipole moment may prove an important design strategy for engineering hybrid improper ferroelectricity.A related strategy is to ask, for a given structure type, what particular distortions need to be added to generate bulk polarisation? Whereas the most common space group for conventional perovskites is Pnma-type) distortions of the perovskite structure. There are several examples of hybrid improper ferroelectric coupling—although not always recognised as such—already present in the literature and we fully anticipate further examples will continue to emerge. Despite the fact that our analysis has been focussed on polarisation in formate perovskite analogues, the general rules developed here extend to all molecular perovskites and to all applications where inversion symmetry breaking is necessary. A recent example is the development of non-linear optics based on dicyanamide perovskite chemistry54.Hence, we can conclude that, from a group-theoretical viewpoint, perovskites with a molecular component are remarkably predisposed to crystallising in polar space groups. This attractive property is a result of the large number of polar coupling schemes generated by distortions accessible to molecular perovskites but inaccessible to conventional inorganic perovskites. We have identified specific combinations of structural distortions that lead to acentric structures and have thereby suggested possible routes for targeted material design. In particular, we highlight explicitly a number of propolar candidates, where the replacement of JT-inactive for JT-active transition-metal cations is likely to drive polarity. We have also found that either A-site quadrupolar order or the activation of columnar shifts—features that are unique to molecular perovskites—can drive polarisation when coupled to conventional , B-site cation at 2af2)3. This was used as an input to the web-based software ISODISTORT55, and the rigid-unit modes and orbital ordering patterns could be identified by inspection.A hypothetical aristotype molecular perovskite was employed as our model system, with the space group s orbital) transform as a singly degenerate irrep, dipoles (p) as triply degenerate, quadrupoles (d) as 5-fold degenerate, etc. To determine the effect on the irreps of lowering the symmetry of SO(3), such as what occurs at any site symmetry in any crystallographic group, we can use descent of symmetry tables. For quadrupoles, if one or more of the irreps enters the symmetric representation (Ag1), then we may consider the action of lowering the point group symmetry to have resulted in multipolar order (and this is how we define quadrupolar order in the context of this paper). In the aristotype ABX3d-orbitals) centred at A/B transform as Eg and Tg2 while at X they transform as Ag1, Bg1, Bg2, Eg. Hence, the problem of determining whether a given irrep of space group Multipoles may be classified as transforming as irreps of the rotational group SO(3). In the absence of any external perturbations to the rotational symmetry , monopoles  in order to mimic electron density in the unit cell at the most general position. We use a setting where the A-site is located at the origin of the perovskite unit cell  orders were considered, a dummy atom was placed on 48All data generated or analysed during this study are included in this published article (and its Supplementary Information files).Supplementary InformationPeer Review File


For all farmers surveyed, there were no significant differences in well-being between farms with and without bTB. For those farms in areas with high bTB prevalence, two of the three measures of subjective well-being showed lower levels of well-being for farmers with a history of bTB (p < 0.05). In conclusion, the paper discusses the policy and methodological implications for future studies of farmer well-being and animal disease.Outbreaks of exotic animal disease, such as Foot and Mouth Disease (FMD) are associated with social and psychological impacts amongst farmers. Whilst claims of similar impacts for endemic diseases have been made, there is little empirical evidence to justify these assertions. This paper provides a descriptive analysis of the social and psychological impacts of bovine Tuberculosis (bTB) in Wales. Specifically, the paper focuses on farmers subjective well-being and presenteeism—their propensity to work suboptimally when suffering mental health problems. Results from longitudinal qualitative interviews with 16 beef and dairy farmers reveal how they derive satisfaction from their work and their emotional connection to animals, whilst the weather and red tape are most likely to affect their quality of life. Data from a postal survey ( Since the 2001 outbreak of Foot and Mouth Disease (FMD) in the United Kingdom, it has become widely accepted that animal disease causes emotional distress to farmers and animal keepers. These socio-psychological impacts arise from the consequences of disease management which disrupt farmers emotional bonds with their animals and the landscapes they create, and the local farming communities in which they are situated . Other sWhilst these accounts focus on the social and psychological impacts associated with emergency responses to exotic diseases, attempts to analyze the impacts of endemic disease are rare. On the face of it, these impacts are arguably likely to be similar: farmers may still experience a sense of loss following the slaughter of infected animals, and economic consequences may cause further mental stress. However, just as with human disease , 15, or The aim of this paper is therefore to analyze the socio-psychological impacts of an endemic cattle disease using measures of subjective well-being (broadly defined as life satisfaction) and “presenteeism” (defined as the extent and effect of attending work whilst ill). To do this, the paper draws on a survey and qualitative interviews of cattle farmers in Wales to assess the impacts of bovine Tuberculosis (bTB). The paper is structured as follows. Firstly, we provide a brief background on bTB before describing in-depth the two measures used to assess the socio-psychological impacts of the disease. Secondly, we analyze farmers' perceptions of well-being. Thirdly, we estimate levels of well-being and presenteeism amongst farmers with and without bTB. Finally, we conclude by considering the wider policy and methodological implications of these findings.It is frequently argued that bTB is the United Kingdom's most pressing endemic animal disease problem, resulting in the slaughter of 55,000 cattle each year at a cost of £100 million to the government , 24. WhiThe socio-psychological impacts of bTB are felt in various ways. The confirmation of a positive bTB test leads to the slaughter of infected animals. Whilst this can trigger emotional impacts to a farmer, and his/her family, the incident may also lead to economic impacts as trade is restricted. Although dairy farmers may continue to sell their milk as normal, selling store cattle or breeding stock is prohibited. The loss of milking cows will impact upon the ability to meet milk supply contracts. Herds under bTB restrictions may be allowed to restock if licenses are agreed by Government vets but negotiating a license may provide additional stress. Moreover, whilst compensation is provided for slaughtered cattle, this is unlikely to cover replacement costs or the indirect costs of additional feed for cattle that would have been sold in normal circumstances. Like FMD, an incident of bTB may also lead to a perception of stigma particularly for those farmers who rely on breeding and showing pedigree animals.Despite these claims, there is no published academic research on the effect of bTB upon farmers' well-being or their ability to work. Some research has highlighted how farmers have become fatalistic, believing there is nothing they can do to prevent a bTB outbreak . Other qPrevious attempts to measure farmers' mental health have relied on objective assessments, such as suicide levels , 32–36 oSubjective well-being (SWB) is derived from the difference between mental well-being  and mental illness  . hedonic dimension includes the subjective experience of happiness or affect and life satisfaction. Hedonic attributes of well-being are those in which the concept of happiness is primarily associated with seeking pleasure and is related to positive affect and having no worries . Secondly, the validated short Warwick Edinburgh Mental Well-being Scale (SWEMWBS) was used (Table 5 for question wording). These statements aim to measure a person's subjective well-being and psychological functioning using eudaimonic and hedonic perspectives. The overall score for this scale is calculated using the total scores from each statement giving a possible range of between 7 and 35. These seven statements have also been used in the UK Household Longitudinal Study and the ONS National Well-being Wheel of Measures . Collins et al.  and presenteeism (SPS) scales. In addition, the survey collected data on respondent characteristics, farm type and management, pressures facing farmers, and attitudes toward bTB policy. Attitudinal questions used 1–5 scales ranging from strongly disagree to strongly agree. Respondents were also able to add additional comments in free-text boxes.n = 582) and the specific areas of Wales that generated the highest proportion of responses were: Carmarthenshire , Pembrokeshire (17%), Mid Powys (12%), and Gwent (10%). Respondents were supplied with a pre-paid return envelope and a reminder was sent to participants 3 weeks into the survey period to encourage response.A postal questionnaire was distributed to 1,800 farmers in Wales in May/June 2013. Respondents were randomly sampled from the Animal and Plant Health Agency's bTB database. The sampling frame was based on a quota sample of 600 each of dairy and beef cattle farmers across Wales, and a randomly selected group of 600 farms that were experiencing a bTB incident at the time survey. The survey was printed in both English and Welsh languages with 8% completing in Welsh which were translated by a native Welsh speaker. The overall response rate was 33%  were used. Multiple response questions were coded into discrete categories. For example, questions asking about farm disease threats were coded into six main categories reflecting different disease types. These included: diseases that affected the fertility of cattle ; bovine Tuberculosis; diseases that affect the productivity of cattle ; diseases that affected youngstock ; other cattle diseases; diseases that affected other animals on the farm ; or none. Each respondent had five opportunities to identify and rank a disease and results from the survey show that in total, 1,680 disease threats were cited (Table 3). A total of 170 farmers (32%) wrote down the maximum of five diseases. Interviews were fully transcribed and analyzed using QSR Nvivo 8 software. Coding was carried out using an inductive approach by initially keeping a record of codes that emerged during transcription alongside those which were anticipated from the existing literature.Survey data were coded and entered into SPSS. Coding followed the instructions relevant to the SWEMWBS and SPS. To test for significant differences between farm types and bTB status, bivariate significance tests  of respondents mentioned this disease as a threat, and it accounted for 26% of all disease mentions. As respondents frequently had more than one enterprise on the farm, diseases of other animals such as sheep, pigs or poultry were also mentioned .When farmers were asked to think about the definition of well-being, what it meant to them and what they thought influenced both positive and negative levels of well-being, analysis of the interviews revealed a number of distinct themes in farmers' understandings of well-being. They are as follows:The main association farmers drew with the concept of well-being was with the idea of health. Farmers distinguished between good physical and mental health as being a sign of well-being. In relation to physical health, farmers commented that being fit and able to work hard was a sign of well-being. For example:I would say well-being to me is being fit and healthy and probably able to do what you want to a reasonable extent. That's how I would describe it.” .“Closely related to physical health was mental health. One farming couple believed that their mental well-being was not a problem because of the enjoyment they received from farming and its way of life. This farmer compared how farming had kept him physically fit compared to others of his age who he had been at school with who had pursued other careers.A second theme was of happiness which connects to the much broader concept of eudaimonia identified in the well-being literature rather than just the physical and mental aspects of health. Several farmers commented that this sense of happiness or contentment came directly from their work even though they were probably working harder than they should. Happiness also stemmed from farmers' work with animals and the sense of pride and enjoyment they got from that. This was particularly the case in relation to pride in farmers stock and being able to complete the cycle of birth to death, or demonstrate success in front of fellow farmers:But I suppose the real joy out of it comes when you're selling something for, you know, good money or taking it to a show and winning or seeing that you've done something with it, like getting a bull into AI or something like that.” .“Connected to the idea of respect, some farmers also commented that being able to get on to do their job was inherently related to well-being. For example:I don't complain about my quality of life, because I'm quite happy, I love my cows, I love my calves, so long as everybody leaves me alone I'm quite happy.” .“Similar ideas are expressed in ideas of good farming  in whichA good network of support and friendship and that sort of thing as well which contributes to a feeling of well-being I would of thought and rather than if you stayed slogging your guts out all day and you don't go through the farm gate you get, you feel probably depressed and down, you get stressed and what have you and you need to have that external factors that help you with feeling good about yourself and life and what you're doing I suppose isn't it I would think” .“Farming is a way of life rather than just an occupation. A comparison is made here to a pleasurable working environment with the example provided below of working in a city or those in other occupations. One farmer felt that there were moments in his working day when he would feel that the location of where he worked counteracted the dull routine jobs particularly during the winter months.It's just been the usual mundane at the moment but there are, I think, now and again a moment or something, it depends where you are on the farm, you look around and you think it's a nice place to be anyway, we've got a few nice spots on the farm where we can view right up to Preselis and so forth and you think it's not a bad spot to be in you know what I mean” .“Farmers were asked to rank in order of importance five farming pressures. Seven broad themes emerged from the coding: finance, the weather, bovine TB, red tape/bureaucracy, paperwork, farm management, and other factors see . It was Weather was cited as the most significant amount of stress by over 70% of farmers. Extreme wind and rain or lack of rain, could all make farming more stressful, placing pressure on budgets by restricting the supply of animal feed. More broadly, farmers connected the weather to seasonality. The seasons influence the kinds of farming activities that are possible, and in doing so impacting upon well-being:I look forward to the cows going out and the grass is growing like hell and I know that the winter chores are coming to an end, because it's just the monotony of scraping out and feeding every day; how these people keep their cows in all year round I don't know because I'm quite happy to see them go out, that's why it's been a pain in the backside the last two summers” .“To me, seeing a calf or a lamb born, seeing new life. I've always been…looked forward to lambing time because the time of year is nice and the days are longer and…I don't know what it is, I don't know, but seeing lambs born and helping them, pulling them whatever, it just gives some…it's a great pleasure.” .“These preferences were individual: some farmers preferred preferred the routine of working during the winter time as the summer months could be very busy times. Harvesting could take up long hours during the day and also was dependable on climatic conditions.Paperwork, form filling and the increased hours farmers have to spend on these tasks rather than physically farming were cited as key influences upon well-being. Whilst this pervaded all aspects of farming, it was also reflected in farmers concerns about the politics of farming and in the Welsh Governments handling of bTB. To demonstrate, one farmer produced a volume of paperwork during the interview:Let me just show you something a minute. That is the total paperwork for one reactor and one doubtful [IR]. Pages and pages [pages rustling]. There is no sense in the amount of paperwork that they [Government] churn out.” .“Financial pressures were connected to the restrictions the weather and bTB placed on the way farmers could manage their farm. For example, the weather affected business profitability due to poor milk prices and higher feed costs. An outbreak of bTB could lead to overstocking requiring more work and employing additional staff to help with the workload. Feed, fertilizer and fuel bills were elevated with more cattle to feed and the lack of income from being unable to sell surplus livestock such as heifers for breeding or store beef cattle unless they went directly to slaughter.Whilst these impacts may appear to play into the cultural identity of “hard work,” they also limited other aspects of “good farming.” For example, in describing what they enjoyed about farming, farmers referred to their contact with animals, participating in agricultural social events, and maintaining a tidy farm:I like to see a tidy farm, I like to see the fields in a nice state and that's really not happening at the moment, I like things to be tidy, the hedges aren't cut, there's ruts in the field…I like improving the farm, I like improving the farm we have done that quite a bit as well, I like putting down concrete and make the road look nice.” .“Whilst this cultural ideal of farming was therefore a strong influence in farmers' well-being, either through regulation, a lack of time or money, pressures such as bTB could prevent these culturally symbolic aspects of farming from happening:What they get frustrated about is that [my children] like showing calves, and they have their favourite calves, that they cannot take them because they're not allowed to go from here. So this year we had to buy them two calves to the other farm, but that means that they have to go every day to the other farm to train them which is five miles away. So it's nowhere near as convenient as doing it here. But the danger with bTB is that it could take their favourite calves away, but I suppose that's part of life. They have to get used to it.” .“7. For the ONS measures, farmers' perceived SWB was lower than for the general population in the UK and Wales as judged by the Annual Population Survey , farmers reported that in the past month they were 75.6% as productive as usual . Farmers were able to accomplish 75.3% of their work compared to their usual level of productivity . They also felt that they were likely to make more mistakes than usual in their work time (26.3%).Farmers' perceptions of presenteeism from the SPS give an overall mean score of 18.78 for respondents to the farmer survey out of a possible range of between 6 and 30. In terms of perceived productivity  than young age groups as judged through the ONS and SPS scales. Well-being did not vary significantly between gender.Differences in farm characteristics and SWB are not statistically significant. For the ONS and SWEMWBS measures, beef farms had the highest levels of perceived well-being and dairy farmers the lowest see , 7. The Farmers who identified finance as a key pressure were statistically significantly more likely to have lower well-being levels for all three measures. These farmers were also more likely to report reductions in the time that they were productive. Farmers reporting red tape as a key pressure were also statistically significantly likely to have lower well-being scores across all three well-being measures.No clear differences were detected between well-being levels and farm bTB status. For the ONS and SPS scales, well-being was marginally higher for bTB-free farms. Results for the SWEMWBS revealed well-being was lower for bTB-free farms. Farmers with bTB were marginally less productive than those without bTB. However, none of these differences were statistically significant, and the small differences suggest there is little difference between farmers' well-being and productivity with and without bTB .p = 0.046), but none of the SWEMWBS measures were significantly different. For the SPS, farmers with bTB were more likely to report that the stresses of their job were hard to handle than those without bTB (p = 0.02).Further analysis compared responses to individual questions in each of the well-being scales to see if there were differences between the individual well-being domains and bTB. On the ONS scale, farmers with bTB were less likely to say that their life was worthwhile . SWEMWBS scores differed by 0.05 and was not statistically significant. Farmers' SPS score on bTB affected farms was 17.94 compared to 19.16 on bTB free farms (p = 0.022). Productivity scores were higher for bTB farms, although these differences were not statistically significant. For farms that have never had bTB, both well-being and productivity scores were higher than for those farms with a bTB history. However, none of these differences were statistically significant whether analysis was conducted for all farms, or separately for those in low and high-risk areas.There are no statistically significant differences between measures of SWB or presenteeism between farmers in areas of different bTB prevalence. However, farmers with bTB in high-risk areas, however, do have statistically significantly lower levels of well-being than those without . For ther = +0.694) for WP2. In other words, farmers with higher levels of SWB were also the most productive. For the same measure, the correlation was lower for bTB-free farms (r = +0.489) and farms that had never had bTB (r = +0.543), indicating lower productivity. This trend was also evident for correlations between productivity measures and the ONS and SWEWBMS. In low risk areas, these correlations show no consistent pattern. In high-risk areas, correlations are stronger for farms with bTB. For example, WP2 is strongly correlated with SPS on farms with bTB when surveyed (r = +0.706) whilst the correlation for bTB free farms was weaker (r = 0.487). For all measures of SWB in high-risk areas, correlations of SWB and productivity were higher for farms with bTB.Measures of SWB were significantly correlated with estimates of productivity for all respondents, and in the low and high-risk areas . For allThese results are, in some senses, unexpected and provide a contrast with previous assessments of the socio-psychological impacts of animal disease. Overall, across three different measures of subjective well-being, results fail to find consistent differences between farms with bTB and those without. Even when examining separate well-being dimensions, there appears to be little difference between well-being in bTB affected farms and those that are not. Furthermore, when considering farmers' assessments of work productivity, results again fail to find statistically significant differences between farms with and without disease. It is only when analysis focuses on areas of high bTB prevalence that there are statistically significant differences in well-being: for both the SPS and ONS measures, levels of well-being amongst farmers with bTB are ~5% lower than those without.Why do the results reported in this paper differ from those for other exotic diseases? One reason might be that diseases such as FMD involve the imposition of emergency regulations that affect farming, its social environment and the natural environment. By contrast, the management of endemic diseases such as bTB can fail to have a clear end in sight, meaning adapting to live with disease diminishes its social impacts. The lack of relationship between well-being and bTB incidence requires further attention. Potentially, this relationship may be mediated by other factors, such as personality, the presence of other diseases and farmers' experiences of the management of bTB. Equally, this analysis does not take into account whether farms have had bTB for a long time and/or lost a large number of animals to bTB. Analysis of the relationship between well-being and other endemic diseases may confirm the findings presented here for bTB. For the moment, however, the evidence presented here suggests that any relationship between bTB and farmer well-being is complex and cannot be reduced to a simplistic conclusion.These results have a number of implications. Firstly, by employing 3 different measures of SWB, we are able to show that assessments of the socio-psychological impacts of animal disease can depend on the precise measure used. Given the extensive testing and validation of the SWEMWBS, it is surprising that results from this scale are not reflected in results for the ONS or SPS scales. One possible explanation is that the abstract and standardized wording for some of the SWEMWBS questions (such as I've been feeling close to people) in comparison to those in the ONS or SPS scales was confusing to farmers. However, piloting of the questionnaire did not reveal any issues. Moreover, questions in all scales addressed similar issues such as feelings of optimism or the ability to deal with problems. Whilst it is not possible from these comparisons to judge which scale is most accurate, evidence from the supporting qualitative research suggests that farmers are affected by incidents of bTB on their farm. What is clear, however, is that the selection of well-being measures to be used in surveys influences results. Researchers are therefore encouraged to employ more than one measure in surveys and use qualitative research to provide methodological triangulation.Secondly, these results may provide some support that farmers can develop coping strategies to displace the impacts of animal disease. Farmers with frequent incidents of bTB may become cynical toward Government policy and fatalistic toward preventing bTB. This lack of self-efficacy may help to mediate the effects of bTB and explain why in some cases, measures of SWB are not even lower for farmers with bTB. For example, a lack of self-efficacy may help to mediate feelings of guilt or failure. Moreover, in framing notions of well-being in relation to cultural concepts of “good farming,” the ability to blame others for failures in disease management allows farmers to retain a sense of self-worth and accomplishment. This presents a conundrum for policy makers: attempts to seek to encourage better biosecurity by aligning it with forms of “good farming” are likely to provide additional threats to farmer well-being if these measures fail. Further research is therefore required to fully understand what coping strategies farmers employ and the precise effects they have upon well-being.Thirdly, that farmers' well-being is associated with the wider spatial epidemiological status is relevant for policy makers and organizations concerned with farmer well-being. These data suggest that those most in need of help and assistance are those farmers with bTB in high-risk areas. The extent of this relationship should be examined further using more detailed epidemiological definitions of bTB risk. In high-risk areas, farmers with bTB may benefit from talking to those without bTB to understand how they cope with the constant threat of bTB, and vice-versa. Alternatively, farmers with bTB may benefit from more targeted well-being interventions to help them manage their farm during a bTB outbreak.Additional research is required to examine how well-being varies amongst age groups and gender. The number of young farmers in our sample is insufficient to draw any conclusions, yet there is no reason why they should not be affected by a bTB incident. Similarly, the role of farming families is worthy of further investigation to examine how other family members may be both affected by bTB and implement coping strategies . FinallyThis paper analyses the impacts of endemic disease on farmer well-being and perceived productivity. Surprisingly, results do not find a clear relationship between measures of well-being and disease incidence. For some measures, farmers with a history of disease in areas at high-risk of disease, do have lower levels of well-being. However, assessments of these impacts will likely depend on the precise well-being measures selected by researchers. As such, there is a continued need for further research, methodological discussion and debate in the study of the socio-psychological impacts of animal disease.The datasets generated for this study will not be made publicly available to ensure confidentiality of respondents.The studies involving human participants were reviewed and approved by School of Planning and Geography Research Ethics Committee. The patients/participants provided their written informed consent to participate in this study.DC conducted original research, analyzed data, and wrote up results. GE contributed to the design of the research methods, analyzed data, and wrote the final draft of the paper.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.


Latent Dirichlet Allocation model to discover the thematic structure of the government’s digital database of parliamentary speeches, and construct from it two-mode networks linking Members of the Parliament to the topics they discuss. Our results show how topic popularity changes over time and allow us to relate the trends followed by political parties in their discourses with specific social, economic and legislative events. Moreover, the community analysis of the two-mode network projections reveals which parties dominate the political debate as well as how much they tend to specialize in a small or large number of topics. Our work demonstrates the benefits of performing quantitative analysis in a domain normally reserved for qualitative approaches, providing an efficient way to measure political activity.Quantitative methods to describe the participation to debate of Members of Parliament and the parties they belong to are lacking. Here we propose a new approach that combines topic modeling with complex networks techniques, and use it to characterize the political discourse at the New Zealand Parliament. We implement a  Topic models have received widespread attention in recent years –4 as theThe New Zealand government has been making parliamentary transcripts (‘Hansard’) available in digital format since 2003. Suitable annotation of these transcripts allow them to be used as a corpus for the development of topic models. This comprehensive corpus of political text can then be examined through a number of lenses. Topic models allow us to monitor the ebb and flow of themes that are discussed in parliament over multiple years and associate particular themes with individual Members of Parliament (MPs). This enables the identification of trends of topics that particular parties follow. That is, we may observe which issues are discussed repeatedly with great interest and which cease to be mentioned.A number of textual analyses of political speeches are concerned with finding where on the political spectrum a speaker falls –8. TopicIn this work we use the parliamentary speeches data, as classified by a Latent Dirichlet Allocation topic model, to construct two-mode networks –12. Thesth century, the left-leaning Labour Party and right-leaning National Party have been the two parties sharing power at New Zealand government. However, in 1996, the method of electing MPs was changed to a mixed-member proportional (MMP) system and these two parties were joined by a number of smaller parties, which sometimes have held the balance of power. In fact, the period analyzed includes a transition of power from the 5th Labour government (1999-2008) to the 5th National government (2008-).For most of the 20This work is of an exploratory nature, in that our goal is twofold: to present a novel quantitative approach of measuring political activity and to demonstrate the benefits of performing quantitative analysis in a domain normally reserved for qualitative approaches, by using a combination of machine learning and complex networks techniques.The remainder of this paper is organized as follows: the Methods and Data sections 2.2 and 2.3 introduce fundamental aspects of topic modeling and two-mode networks respectively and outline the preparation and organization of our data; Sections 3 and 4 present the results of our analyses alongside a discussion and our conclusions.The semantic data we are using for our analyses are extracted from the New Zealand Hansard database . HansardShop Trading Hours Amendment Bill and was published in Hansard Volume 716 on the 17th of August 2016 [Once these data are obtained, we observe that many speeches are rather short and contain little topical content. An example is given below, which comes from a committee discussion on the ust 2016 :CHAIRPERSON (Lindsay Tisch): Just a point: this debate concludes at quarter past and to whoever is speaking at the time, I will be stopping it at that point.”“In an attempt to remove these non-topical speeches, we have removed from our database speeches with 150 words or less, which constitute about 20% of the database. This cut-off is shown in D and the total number of topics as K. Additionally, the order of words that comprise the document is not considered, only the frequency with which words appear.The process of topic modeling involves utilizing a set of algorithms that have been developed to understand the underlying thematic structure of a corpus. The simplest and most commonly used topic model is Latent Dirichlet Allocation (LDA) . Within hidden variables. The word-distributions per document are observed, while the topic structure—per-word topic assignment, per-document topic proportions and per-corpus topic distributions—are hidden elements. Therefore, the central computational problem for LDA is to infer the hidden structure that likely generated the observed corpus. This means computing the conditional distribution of the hidden variables given what is observed. This conditional distribution is usually referred to as the posterior and can be expressed asβK1: are all topics in the corpus, θD1: are the per-corpus topic proportions, zD1: the per-corpus topic assignments and wD1: the whole set of observed words. Unfortunately, computing the posterior is computationally unfeasible and hence needs to be approximated by an inference algorithm. Consequently, topic modeling algorithms are commonly classified as sampling-based algorithms or variational algorithms [From the perspective of LDA, documents are imagined to be the result of a generative process. This is the process by which the model assumes the documents arose given certain gorithms .In this work, we used the MALLET software package  for the LDA assumes the topics are the same for all documents, and only the topic proportions vary. Therefore, MALLET requires an input which specifies the number of topics to be discovered. Choosing this number is critical to the success of a topic model, as too few topics may merge distinct themes, while too many topics may introduce many “themes” consisting of vocabularies that appear to have nothing in common, or even start splitting topics that were identifiable at smaller input values. For our analysis it is important that the topics are easily identifiable and distinct from one another. We found that 30 topics satisfies these requirements. Identified topics and their corresponding keywords can be found in G = {U, V, E}, where U and V are disjoint sets of nodes and E = {: ui ∈ U, vj ∈ V} is the set of links connecting these nodes. For our purposes, the sets U and V correspond to the sets of MPs and topics, and the set E represents the links that emerge when an MP speaks sufficiently frequently about a topic. No connections among nodes of the same set are allowed in the bipartite network, that is, MPs are connected only to topics not other MPs and vice versa. Each set of nodes can have independent properties, such as the degree distributions, or the number of nodes (system size).A great number of real world networks are, in fact, one-mode projections of bipartite architectures comprised of two different types of nodes. Such architectures are called two-mode networks, also known as bipartite networks. A two-mode network is, therefore, mathematically defined as a bipartite graph Once we find the set of topics that a particular MP speaks about ‘often’ enough (this criterion is defined below), these are represented as links between the MP and those topics. After this process is completed for all MPs, we can construct a two-mode network where nodes representing MPs are connected only to nodes representing topics, and vice versa.projection of the original two-mode network.Bipartite structures play an important role in the analysis of social and economic networks. They are normally used to represent conceptual relations—such as membership, affiliation, collaboration, employment, ownership and others—between two different types of entities within a system –12. Oftesimple weighting method [Topic modeling results in a natural bipartite network with projections that can be easily interpreted. The projections of a bipartite network are obtained by connecting nodes which share a common neighbor. That is, if two MPs are both linked to the same topic in the bipartite network, then they are linked in the MP-projection. For a bipartite network, this process results in two completely separated components, each composed exclusively of one type of node (MPs and topics in our case). g method , wherebycomplete subgraphs (where every node is connected to every other node in the subgraph) of MPs are generated by every topic. This means that every MP that speaks about a popular topic is connected to every other MP that speaks about that same topic. The existence of popular topics can make analyses such as community detection challenging in the absence of weighting.The weighting in these projections offers a way to eliminate edges that represent tenuous links. This is important, as 4 words in the entire term were removed form the network for the lack of significance in the volume of words spoken.In order to build bipartite networks connecting MPs to topics, we looked at the corpus of each MPs speeches in more detail. We considered an MP to be connected to a topic when at least 6.7% of the MP’s speeches over the course of a year was assigned to that topic by MALLET. This occurs when MPs talk about a topic twice as much as would be expected if they were talking about all topics equally within a year. This method, removes topics that MPs only touch on briefly or in passing, which does not indicate engagement with the topic. Finally, MPs that had spoken less than 10Despite having fewer MPs, opposition parties tend to have a greater total word count than the governing party. Allowing a decomposition by party, we ran a topic model on data concatenated by MP and year. The topic proportions obtained over a total number of 30 topics are normalized for each year so that they are comparable across a time span of 14 years. Proceeding this way, we can reproduce the evolution of topic popularity over time at the Parliament and its decomposition for each of the three most represented parties. Clear trends and differences across parties are visible in Figs Changes in topics being discussed often correlate with real world events such as the financial crisis of 2008 , Canterbth to 50th parliaments resulting from the process described above are shown in The MP-projected networks for the 47Before examining the structure of the network, it is important to note that in all networks a number of MPs are not connected to any others. This is a reflection of our methodology. The 6.7% threshold that is applied to filter topics may in fact remove all topics in the unlikely scenario that the MP in question talks about many topics in roughly equal proportion. MPs with diverse interests may not be identified as fitting into any particular community.A striking difference between the MP networks of the first two parliaments and last two is the party composition of visible communities. In the first two  the communities are quite party-wise diverse, while in the last two  there is a close-knit community made up of National Party MPs. This is corroborated by the three largest communities composition, for every term in our analysis, shown in th parliament, since it has more MPs speaking about three topics than in the previous parliaments.The degree distributions in th and 50th parliaments, the degree distributions of the MPs networks become more uniform. The tendency towards more uniformly connected networks suggests that National MPs (which are the most abundant during the last terms) share more common interests among them than Labour party MPs did when they were in government. With the aid of th and 50th parliaments show the formation of a well-defined National Party group. In particular, in the 50th parliament, the members of the largest community are connected to at least 45 other MPs, most of whom are also members of this tightly interconnected National Party group.From Also supporting this idea of a close-knit community made up of National Party MPs is Topic models provide a way to parse human speech and extract themes from large bodies of text that are often difficult and time consuming to analyze manually. In few cases is it more important to gather and process this information than in the speeches of those people that control the legislative and political direction of a country. Topic modeling is unlikely to replace traditional media analysis of political speech, however, here we have shown that it can be a useful tool for examining the themes and trends in political discourse from a quantitative perspective.Super City via the amalgamation of numerous smaller councils  or because the MP had spoken too few words (below ten thousand words) during the entire term.(PDF)Click here for additional data file.S2 TableTopics identified and their respective key words from Mallet. *Topics that were removed from the networks. **Sometimes just part of Māori words appear due to accent marks. Those are not readable in the format used in Mallet (e.g. ‘whānau’ would appear as ‘wh’ only). Speeches in Māori are translated to English, i.e. the corpus in this work contains these speeches in their translated versions. Therefore, this topic needed to be removed from the network.(PDF)Click here for additional data file.S1 FigTime evolution of topic proportions, as discussed at Parliament and its decomposition by party.(EPS)Click here for additional data file.S2 FigFrequency of number of words spoken by MP during the four parliaments.(EPS)Click here for additional data file.


There are errors in the Author Contributions. The correct contributions are:Conceptualization: Marie-Hélène MoncelData curation: Alison Pereira, Sébastien Nomade, Jean-Jacques Bahain, Pierre Voinchet, Marcello PipernoFormal analysis: Marie-Hélène Moncel, Carmen Santagata, Alison Pereira, Sébastien Nomade, Jean-Jacques Bahain, Pierre VoinchetFunding acquisition: Marie-Hélène MoncelInvestigation: Marie-Hélène Moncel, Carmen Santagata, Alison Pereira, Sébastien Nomade, Jean-Jacques Bahain, Pierre VoinchetMethodology: Marie-Hélène Moncel, Sébastien Nomade, Jean-Jacques Bahain, Pierre VoinchetSupervision: Marie-Hélène MoncelValidation: Marie-Hélène MoncelWriting–original draft: Marie-Hélène Moncel, Alison Pereira, Sébastien Nomade, Jean-Jacques Bahain, Pierre VoinchetWriting–review & editing: Marie-Hélène Moncel


Polyvinylidene Fluoride (PVDF) piezoelectric electrospun nanofibers have been intensively used for sensing and actuation applications in the last decade. However, in most cases, random PVDF piezoelectric nanofiber mats have moderate piezoelectric response compared to aligned PVDF nanofibers. In this work, we demonstrate the effect of alignment conducted by a collector setup composed of two-metal bars with gab inside where the aligned fiber can be formed. That is what we called static aligned nanofibers, which is distinct from the dynamic traditional technique using a high speed rotating drum. The two-bar system shows a superior alignment degree for the PVDF nanofibers. Also, the effect of added carbon nanotubes (CNTs) of different concentrations to PVDF nanofibers is studied to observe the enhancement of piezoelectric response of PVDF nanofibers. Improvement of β-phase content of aligned (PVDF) nanofibers, as compared to randomly orientated fibers, is achieved. Significant change in the piezoelectricity of PVDF fiber is produced with added CNTs with saturation response in the case of 0.3 wt % doping of CNTs, and piezoelectric sensitivity of 73.8 mV/g with applied masses down to 100 g. The polarization is caused by a reorientation of the net dipole moment of polymer polar groups into a particular direction ,2,3,4,5.γ phases . If the γ phases ,9,10,11.Electrospinning is one of the most popular techniques for nanofiber (NF) fabrication ,13,14,15The impact of the addition of carbon nanotubes (CNTs) on the crystal structure and piezoelectric properties of PVDF nanofiber has been widely studied. PVDF/CNTs nanofiber composite presents high β phase formation, high electrical signal, and improved mechanical properties that make CNTs nanofiber composite a good candidate for a wide range of sensing applications. The increase of linear behavior in output voltages of piezoelectric signals for aligned and randomly-oriented electrospun PVDF/CNTs membranes was investigated with applied loading up to 350 N, and showed high stability during cyclic dynamic compression and tension .Several techniques have been investigated to increase the degree of alignment of polymeric nanofiber through electrospinning ; continu3-0.22 SrTiO3)/PVDF-TrFE composites were fabricated by the electrospinning method, observing the effect of the drum collector rotation speed from 0 to 1500 rpm on the degree of alignment of electrospun nanofiber. An increase of nanofiber orientation resulted that increased the piezoelectric output [Electrospun piezoelectric polymers have the potential to be used in various applications ,25,26,27c output . Air prec output . Simulatc output .In this study, Piezoelectric PVDF/CNTs nanofibers were fabricated through electrospinning. Aligned fiber through 2-metal bars was investigated by studying the effect of the alignment and CNT concentration on the piezoelectric response. This alignment technique is static and alternates the traditional rotating drum alignment procedure. This is important for further study of scaling-up the generated nanofibers mat. The fabricated mats of the collected aligned and random nanofibers were analyzed by scanning electron microscope (SEM), and the β phase content was calculated by Fourier transform (FT-IR) analysis. The crystal phase of the piezoelectric nanofibers was investigated by X-ray diffraction (XRD) analysis ; COMSOL The main precursor of Polyvinylidene Fluoride (PVDF)  is supplied by ARKEMA. Multi-walled carbon nanotubes (CNTs), from Cheaptubes Inc.  are added within different weight percentage. The outer diameter of these tubes is in the range of 10–20 nm, and the inner diameter between 3–5 nm.Electrospinning was performed by adding 20 mL of dimethyl formamide (DMF)  solvent into a 3 gm polyvinylidene difluoride (PVDF) to get 15 wt % PVDF concentration. A plastic syringe tipped with a stainless steel needle was filled with 3 mL of the PVDF solution. The positive voltages came from a high voltage supply CZE1000R  to the metal needle, for application of bias values around (25 kV) with constant rate of (1.5 mL/h) using a syringe pump NE1000  with needle-to-collector distance of 10 cm. Random PVDF nanofibers were obtained using a normal metal plate collector covered with aluminum foil which was connected to ground. For comparison, Aligned PVDF fiber was fabricated by 2-metal bars of 1.5 cm length, as shown in Ii is the length of the fiber, Itot is the total sum of fiber lengths; θi is the angle between each fiber and the x-axis.Nanofibers are formed between the 2-bars due to the high electric field. We introduced a finite element simulation for the proposed collectors during the study, showing both electric and electric field vectors between the 2 bars. The finite element package COMSOL Multiphysics® Version 5  was used to demonstrate the fields profiles. The simulations were investigated using 3D module. SEM images were analyzed using Fourier Transform through the usage of MATLAB algorithm, Image-Based Fiber Orientation Calculator by university of Minnesota-Academic Use License was obtained , and imaα), where α = 0 for complete randomness of the nanofibers mats and α = 1 for completely aligned nanofibers. Anisotropy index is calculated from an equation relates the eigenvalues , of the orientation matrix defined as follows:The alignment scheme is determined through a factor called anisotropy index (The morphology of PVDF NFs was observed by a Scanning Electron Microscope (JEOL JSM-6010LV-SEM). The diameter of NFs was analyzed using (Image J) software. The crystal phase of NFs was measured with X-ray Diffractometer (XRD) , and Fourier Transform Infra Red Spectrometer (FT-IR)  was used for β phase content calculation. Simulation for alignment setup with orientation of electric field directions for plate and bars collectors was introduced using COMSOL finite-eliminate software. Degree of NFs alignment and anisotropy was observed using image-based fiber orientation and alignment calculator software program which is based on Fourier transform methods. Piezoelectric properties of the nanofiber mats are measured through a simple setup. The nanofiber mats of dimensions 2 cm × 2 cm were placed between two foil sheets, and pressed by different applied weights. Then, the generated voltage was detected through two connected shielded wires, pasted on the foil sheets, to a high impedance oscilloscope; Tektronix 3012 .To show the electric field profiles, COMSOL Multiphysics was used to analyze the two collectors, to show the field profile on the normal collector and the 2 bars collector. It can be observed that, in the conventional collector case, the electric field profile and vectors are not oriented in a certain direction; however, in the two-bar collector, the electric field vectors are forming stretching forces in the gap between the two-bar. This can explain the differences in the fibers’ common oriented direction. Surface morphology is shown in Degree of alignment for the nanofibers mats  to prove the two-bar collector effect on the PVDF nanofibers alignment. The orientation matrix for the two-bar collector and conventional collector was obtained. Degree of alignment for the PVDF/Carbon nanotubes (CNTs) is calculated for two samples with 0.1% and 0.3% CNTs. The anisotropy index still shows an improvement in the two-bars setup for PVDF/CNTs samples when compared to conventional collector; however, the alignment percentage decreased due to the presence of CNTs. This may be explained by the effect of CNTs charge on the orientation of electrospun nanofiber. θ = 20.6°) increased for the aligned nanofibers with two-bars as compared to random nanofiber, whereas the non-polar α-phase (2θ = 40°) exhibited the opposite trend , 766 cm−1 , and 795 cm−1 (CH2 rocking) were due to the PVDF α-phase, while the vibrational band at 840 cm−1, 879 cm−1 and 1270 cm−1 corresponded to the PVDF β-phase (F(β), represents the β phase content, and Aα and Aβ are the absorbance at 766 and 840 cm−1 respectively. By calculating the previous equation according to the obtained IR result, the β-phase content for the aligned NF was 86%, whereas the random NFs exhibited β-phase contents of 74.2%.The IR vibrational bands observed at 612 cm β-phase . AccordiRegarding the piezoelectric analysis, each mat was exposed to pressures from different masses, and the generated peak-to-peak voltages were recorded. Both The work demonstrates the ability to generate highly-aligned PVDF nanofibers through static-alignment of the collector side of electrospinning setup. The synthesized PVDF nanofibers are studied with and without added in-situ CNTs. The two-bar collector system shows a great enhancement in the alignment degree for the PVDF nanofibers. Our results show that both alignment and added CNTs increased the formation of beta sheets inside the nanofibers, which correlates to a better polarizability inside the material. Consequently, the piezoelectric sensitivity of alignment PVDF doped with 0.3 wt % of CNTs was improved up to the range of 73.8 mV/g when applying weights up to 100 g, where the generated nanofibers mat can be used as a piezoelectric sensor. At higher applied weights, our synthesized nanofibers show saturation behavior which leads to the possibility of using our nanofibers as an electronic sensor with a steady output of generated voltage.


In this study, two zeolitic imidazolate frameworks (ZIFs) called ZIF-4 and ZIF-zni (zni is the network topology) were characterized by sorption studies regarding their paraffin/olefin separation potential. In particular, equilibrated pure and mixed gas adsorption isotherms of ethane and ethene were recorded at 293 K up to 3 MPa. ZIF-4 exhibits selectivities for ethane in the range of 1.5–3, which is promising for continuous pressure swing adsorption (PSA). ZIF-4 shows high cycle stability with promising separation potential regarding ethane, which results in purification of the more industrial desired olefin. Furthermore, both ZIF materials were implemented in Matrimid to prepare a mixed matrix membrane (MMM) and were used in the continuous separation of a propane/propene mixture. The separation performance of the neat polymer is drastically increased after embedding porous ZIF-4 crystals in the Matrimid matrix, especially at higher feed pressures (3–5 barg). Due to the smaller kinetic diameter of the olefin, the permeability is higher compared to the paraffin. In 2(BTC)3 ,7,8. The2(BTC)3 ,10. A se2(BTC)3 , ZIF-8 [2(BTC)3 , UTSA-102(BTC)3 , and PCN2(BTC)3  and seve2(BTC)3 ,16,17. HZIF-4 belongs to the group of so-called zeolitic imidazolate frameworks (ZIF) that consists of zinc metal centers connected to imidazole-based organic linker molecules. Besides ZIF-4, which possesses cag topology, different polymorphs such as ZIF-zni (zni topology), ZIF-1 (BCT), ZIF-3 (DFT), ZIF-6 (GIS), and ZIF-10 (MER) are known . From thTo improve the separation performance in breakthrough and pressure swing adsorption (PSA) experiments, high-pressure isotherms are beneficial to determine the optimal sorption conditions for dynamic separation. Additionally, a volumetric set-up combined with a gravimetric system allowed us to determine sorption equilibria of binary gas mixtures of ethane and ethene. From these results, we were able to derive the equilibrium selectivity for this sorption process. 2 capture [To prove the separation concept with porous materials, continuous separation experiments are indispensable. There are three different instrument-based solutions: the PSA setup, used, e.g., for air separation  and hydr capture ,23, hydr capture ,25, and  capture . The advThis also applies to membrane-based technologies. The process can be described as a transport across the membrane due to a concentration gradient (Fick’s law). However, for dense (polymer-based) membranes, which transport gas molecules by the solution-diffusion mechanism, the amount adsorbed depends strongly on the ability of the gas to be dissolved in the polymeric matrix, which can be approximated via the condensability of the gas or its molecular size . Mixed mIn this study, the continuous separation potential of ZIF-4 powder for an ethane/ethene gas mixture was tested in a commercial PSA setup. To find the best separation conditions for the continuous setup, high-pressure and mixed adsorption isotherms for ethane and ethene have been recorded. In addition, MMMs with implemented ZIF-4 and ZIF-zni crystals were investigated with respect to their separation potential for propane/propene gas mixtures, which could not be separated using ZIF-4 powder as described in our previous paper [The equilibrium adsorption isotherms of pure ethane and ethene at 20 °C up to 3 MPa  show thep > 500 kPa). This implies that the best separation results should be obtained up to a pressure of about 100 kPa. To demonstrate the ideal behavior for this mixture, we measured the mixed gas adsorption isotherm (The presented data show a higher sorption capacity for ethane compared to ethene at lower pressure (0–300 kPa). However, at higher pressure (300–3000 kPa), the loading of ethane reaches a saturation plateau while the loading of ethene still increases. This leads to a decrease in selectivity until a slightly higher loading is found for ethene compared to ethane  based on the Toth fits from single gas isotherms. Because of the good agreement, ideal adsorption behavior can be assumed. From the plot, partial loadings for a certain gas phase fraction can be obtained. In α = 1.24 regarding ethane at 301 K and 138 kPa for an ethane/ethene mixture with a ratio of 28/72%. The AC Nuxit-Al has a nearly constant selectivity of α = 1.49–1.56 over the entire partial pressure range of ethane at 293 K and 101 kPa. The experimentally observed ethane selectivities for ZIF-4 range from 1.5 to 3, which is higher than the values reported for active carbons. In contrast, zeolites typically exhibit a preferential sorption for ethene as a consequence of favorable adsorption sites for unsaturated hydrocarbons [α = 4.7–7.4 for the sorption of ethene over ethane on zeolite 13X at 323 K and 138 kPa. Thus, ZIF-4 appears to be a promising adsorbent resulting in a higher retention of ethane for PSA experiments aiming at the continuous separation of olefin/paraffin mixtures. The preferred adsorption of ethane in the absence of strong adsorption sites is triggered by the different critical temperatures of the gases. Sakurovs et al. [In the literature, mixture data of ethane and ethene on common adsorbents like activated carbons  with low amounts of ethane (closed cycles) are displayed. For clarity, all data points are connected and a section of the complete measurements is added cf. , inset. In the insert, the switch between both columns is pictured by the concentration drop of ethene. The relative concentration of each peak is related to the degree of dilution by the purge gas nitrogen. By neglecting the amount of nitrogen in the product, the purity of ethene amounts to 99.16%. This renders ZIF-4 as beneficial adsorbents for continuous ethene purification in order to obtain a high level of purity. With the low operation temperature and no need to employ high pressure, the running costs are expected to be low. Therefore, the utilization of ZIF-4 in PSA systems seems to be an economic alternative to ethene purification by low temperature distillation. The characterization of the applied MMMs regarding the layer thickness and the embedding of the ZIF-4 and ZIF-zni crystals was performed with a scanning electron microscope. The SEM pictures of the MMM cross sections in low and higher magnification are shown in The images in propene/propane) for both gases on MMMs with ZIF-4, ZIF-zni and neat Matrimid is displayed in A feed mixture of propane and propene was used to investigate the influence of the porous adsorbents ZIF-4 and ZIF-zni as filler in MMMs in comparison to the neat polymer material Matrimid. The measured permeability as well as the calculated separation factor . Under these conditions, both the propene permeability and the separation factor are almost doubled in comparison with the neat Matrimid. When adding the denser phase ZIF-zni, the separation factor is slightly enhanced in comparison to the neat Matrimid (1.5 to ≈2.3). In contrary to this, the propene permeability is almost six times higher compared to the neat polymer film. These high permeabilities are most likely a consequence of the void-rich ZIF-zni MMM, since the contact between the filler and the polymer phase is weak. The permeation results are in complete accordance with the SEM results in ZIF-4 was synthesized as described elsewhere [The synthesis for ZIF-zni is based on the pathway of Bennett et al.  with sma2O3 support with a pore diameter of 1.8 µm. The hardening procedure occurs under a DCM atmosphere. After evaporation of the solvent in Petri dishes, the MMMs were activated in the measurement cell at 150 °C and under a nitrogen flow to remove residual solvent . For pictures of cross sections by scanning electron microscopy , the MMMs were broken into two pieces and fixed on a brass support with carbon pads. Subsequently, they were sputtered with carbon for electric conductivity.Prior to the preparation of the mixed matrix membranes, the required ZIF-4 and ZIF-zni crystals were synthesized as described in The pure gas equilibrium isotherms of ethane and ethene at 20 °C were measured with a gravimetric-magnetic suspension balance (MSB) from Rubotherm, Bochum, Germany (resolution 10 µg). For the measurements, 0.3 g of ZIF-4 were activated at 200 °C for at least 12 h in vacuum. The respective gas was dosed into the sample chamber. The weight as well as the gas pressure was recorded. The equilibrium state was assumed when the weight change was <10 µg and pressure was constant for 10 min. For further measurement points, the gas pressure was stepwise elevated.For the mixed gas equilibrium isotherm, a manometric-gravimetric setup was employed. This is assembled by an MSB, which is integrated in a manometric setup see .The gas phase composition that is necessary for balancing the manometry was analyzed with the gas chromatograph HP5890 series II from Hewlett-Packard equipped with a flame ionization detector (FID). For the measurement, ethane was first submitted to the measuring chamber until equilibrium (weight change <10 µg and constant pressure for 10 min) was reached at 100 kPa. Subsequently, portions of ethene were dosed stepwise at 100 kPa total pressure to reduce the amount of ethane in the gas phase and to enforce a partial desorption of ethane from ZIF-4. The equilibrium of the mixed gases has been calculated from the mass balance given by pairs of pressure, temperature, gas phase composition, and loading before and after the dosing of ethene to the measuring chamber. For mixed gas adsorption measurements, an amount of 1.5 g of activated ZIF-4 was used.The selectivity was calculated from experimental data cf.  by usingThe selectivity for activated carbons and zeolites are estimated from a set of experimental data taken from the literature using the same equation. We have added this information to the experimental section.For the continuous separation of ethane and ethene mixtures, the lab scale pressure swing adsorption setup PSA-300LC from L&C Science and Technology, Hialeah, FL, USA) was used. A thermostat with Peltier element was employed to keep a constant temperature of 20 °C in both adsorber columns . These are filled with 1.164 g (column 1) and 1.188 g (column 2) of ZIF-4 with a particle size between 106 and 250 µm. Both packings were ex situ activated in a vacuum oven at 150 °C for 20 h. The inner volume of one column is ca. 2.21 cm³. For continuous separation, an operation mode similar to the Skarstrom cycle ,40 was uOne cycle consists of the adsorption and purging step of the first column and the purging and adsorption step of the second column. The adsorption step of the first column runs parallel to the purging step of the second column. This cycle can be repeated ad libitum and defines the length of the separation experiment. N/min), an equimolar mixture of ethane and ethene, is loaded and separated during the adsorption step. For the purging step, nitrogen (N/min) was used to regenerate the loaded column in counter current to the feed. The feed . This expression is related to the process described by Rao et al. for separating azeotropic liquid mixtures where a third component was used for purging . The cycN/min) is transported to the membrane in the permeation cell. The molecules passing the membrane are called permeate and the retained molecules leave the permeation cell as retentate. To preserve a high driving force and to enable analysis, the permeate is swept with nitrogen (N/min) toward a gas chromatograph . The permeation experiment was carried out at a constant temperature of 150 °C and at different feed pressures . Measurements were carried out until the equilibrium was reached, which was usually achieved after 24 h of continuous testing.For the permeation measurements, a home-built apparatus , the separation performance is reduced due to the smaller difference in loading for both gases. Finally, the loading of ethene exceeds the values of ethane, which implies a higher retention of the requested gas and is adverse for its purification.The mixed isotherms for ethane and ethene showed ideal behavior, which allows to predict further equilibrium data for mixtures with ideal adsorption solution theory based on pure component isotherms. The continuous separation of equimolar ethane/ethene mixtures was successfully demonstrated with ZIF-4 powder in the lab scale pressure swing adsorption setup. Over a specific period of 50 h, the separation performance does not decline, which is related to the stable adsorption behavior of ZIF-4 and the proper regeneration during the purging step. Therefore, the separation is more influenced by static adsorption than kinetic effects. The embedding of ZIF-4 and ZIF-zni crystals in the polymer matrix Matrimid was successful and showed beneficial properties of the resulting MMM with respect to the permeability of propane and propene compared to the neat Matrimid membrane. The best separation factor of propane and propene was achieved with ZIF-4 crystals in MMMs that doubled the value of the neat Matrimid. Separation capabilities might be further improved in future studies by increasing the interaction between the filler and the matrix.


Long non-coding RNAs (lncRNAs) have been widely known to have an appreciable effect in physiology and pathology. In tooth regeneration, periodontal ligament stem cells (PDLSCs) are regarded as a key effector, whereas, how lncRNA acts in the osteogenic differentiation of PDLSCs have not been completely understood. This study aims to find out the relationship between lncRNA DANCR and the proliferation and osteogenic differentiation of PDLSCs.Microarray was used to observe the different expression of lncRNAs in differentiated and undifferentiated PDLSCs. And then osteogenic-related lncRNA, DANCR was screened out. Its effects on proliferation and osteogenic differentiation was explored by constructing an overexpression and inhibition model. qRT-PCR was used to detect the mRNA expression of osteogenesis related genes. MTT assay was performed to assess the effects of DANCR on cell growth curve. To quantify the effects of DANCR on osteogenic differentiation of PDLSCs, ALP staining and alizarin red was performed in basic culture medium and osteogenic medium. Data were statistically processed.Compared with the undifferentiated PDLSCs, the alizarin red staining level was higher in differentiated PDLSCs. And the expressions of osteogenic differentiation marker genes Runt-related transcription factor 2 (Runx2), osteocalcin (OCN) and bone morphogenetic protein (BMP-2) were significantly increased in the differentiated PDLSCs. Furthermore, we noticed that comparing with control groups, the expression of lncRNA DANCR decreases markedly in osteogenically induced PDLSCs. DANCR promoted proliferation of PDLSCs, as evidenced by cell viability. Further investigation has proven that the downregulation of DANCR shows in the calcium sediment forming, alkaline phosphatase (ALP) activation and some osteogenic-related gene markers’ upregulation including Runx2, OCN and BMP-2, which finally results in the osteogenic differentiation of PDLSCs following the transfection and induction. Conversely, DANCR upregulation was shown to repress the osteogenic differentiation potential of PDLSCs.The osteogenic differentiation of PDLSCs has proven to related to the down regulation of lncRNA DANCR. And this paper throws light on the effects of DANCR in the process of PDLSCs’ osteogenic differentiation. Dental problems caused by dental caries, periodontal disease and tooth injury compromise the oral and general health issues . PeriodoThe periodontium is composed of periodontal ligament (PDL) tissue, alveolar bone, cementum and gingival tissue . PeriodoLong non-coding RNAs (lncRNAs), which cannot code protein and are more than 200 nt, play crucial roles in both cell fate determination and disease pathogenesis . PreviouLong non-coding RNA differentiation antagonizing non-protein coding RNA (LncRNA DANCR) was the first lncRNA demonstrated to have relationship with the suppression of progenitor differentiation . The funWe currently focus on the effects of lncRNA DANCR on PDLSCs’ osteogenic differentiation. We showed that lncRNA DANCR was significantly downregulated in osteogenically induced PDLSCs compared to that undifferentiated counterparts. Further examination indicated that the inhibition of DANCR was positively related to the PDLSCs’ osteogenic differentiation following the transfection and induction. However, DANCR upregulation was shown to repress PDLSCs’ osteogenic differentiation. These data suggest that lncRNA DANCR get involved in cell osteogenic differentiation of PDLSCs.The alizarin red staining was performed to examine the osteogenic differentiation of PDLSCs. As showed in Fig. After transfection, the result of qRT-PCR showed that the expression of lncRNA DNACR was significantly increased in pcDNA3.1-DANCR group while it was notably decreased in pCMV-sh-DANCR group Fig. a. The reWe have previously demonstrated that the level of DANCR was decreased in PDLSC osteogenic differentiation. Based on this finding, we sought to further investigate the effects of DANCR in the osteoblastic differentiation of PDLSCs. As showed in Fig. In this study, we aimed at finding the role of lncRNA DANCR in the osteogenic differentiation after transfection of PDLSCs. MTT assay was performed to measure the cell proliferation, and the results indicated that cell proliferation was suppressed in pCMV-sh-DANCR group and was enhanced in pcDNA3.1-DANCR group. And the result of qRT-PCR showed that the osteogenic differentiation marker genes, including Runx2, OCN, and BMP-2, were significantly increased in pCMV-sh-DANCR group. We performed ALP activity and alizarin red staining  assays tRecently, many studies have focused on the functional deficiency of stem cells in several disease conditions. PDLSCs are mesenchymal origin that can differentiate into cementum-forming cells and osteoblasts and thus have been regarded as the most important seed cells for periodontal tissue regeneration .Through Besides, there are some reports similar to our study. Zhang et al. found that lncRNA DANCR was downregulatedin human bone marrow-derived mesenchymal stem cells (HBMSCs) during osteogenic differentiation, and transfection with si-DANCR significantly enhanced the cell viability and promoted osteogenic differentiation of HBMSCs . And JiaMoreover, the molecular mechanism of this differentiation potency is poorly understood. In the study of QU et al., pathway analysis suggested that several kinds of signaling pathways were involved in the osteogenic differentiation process in hPDLSCs, including TGF-beta signaling pathway, Toll-like receptor signaling pathway, MAPK signaling pathway and osteoclast differentiation, etc. . And theAt present, lncRNA research has been widely used in the study of osteogenic differentiation, Yang et al. found that lncRNA Reg1cp abnormal mutation can lead to human high bone mass syndrome . The rolDown-regulated of lncRNA DANCR promotes osteogenic differentiation of PDLSCs, which maybe a novel therapeutic strategy that can be used for the dental tissues.The periodontal ligament (PDL) was collected from patients in the Shanghai East Hospital, Shanghai Jiao Tong University School of Medicine. The first or the second premolar teeth were respectively collected from six healthy patients ranging from 18 to 25 years old for orthodontic purposes. This study was approved by the ethics committee of Shanghai East Hospital Affiliated to Tongji University. Written informed consent was obtained from all participants at the beginning of the study.. All experiments were performed in accordance with approved guidelines and regulations from Shanghai East Hospital Affiliated to Tongji University of Medicine Ethics Committee. In this study, the experimental data analysts applied the blind method for data analysis and statistics to avoid deviations in the data processing process.3. Then, the PDL tissue was digested in 3 mg/ml collagenase I  and dispersed for 45 min at 37 °C. Then, added an equal volume of medium to terminate the digestion. The solution was centrifuge d at 1000 r/min for 5 min, and the supernatant was discarded. Next, the tissues were resuspended in 1 ml of medium and inoculated into 6-well plates and cultured in the environment of 37 °C and 5% CO2. After every 2 or 3 days, the medium would be changed until the cells appeared from the edge of the tissue after 3–8 days. Then the cells were cultured as Zhang et al. reported and four generations of poly-clonal PDLSCs were used in this study [The extracted teeth were cultured in 90% Dulbecco’s Modified Eagle Medium (DMEM) with 100 u/ml penicillin, 100 μg/ml streptomycin, and 10% fetal bovine serum (FBS). The PDL tissue was obtained by scraping the root surface of teeth after washing 6–8 times with PBS (containing penicillin/streptomycin). The centrifugation of 5 min in 1000 r/min was performed for PDL tissue which was cut into small pieces of 1 mm5 cells. Osteogenic differentiation induction medium , 10 mmol/L of beta-glycerophosphate  and 10 ng/mL of dexamethasone diluted in 10% FBSa-MEM) was given when cells reached 80% confluence. After every 3 days, the conditioned medium was changed. Cells cultured with standard culturing medium were designated as control group. After 21 days of culturing, the differentiated PDLSCs were formed.PDLSCs were transferred into a 6 well plate, and per well has the density of 1 × 106 cells, were prepared, and TRIzol reagent  was used to isolate total RNA. And the RNA was amplified and transcribed into fluorescent cDNA. Human LncRNA Array was used to hybridize with labeled samples. Differentially expressed lncRNAs were identified through Volcano Plot filtering with the threshold of fold change > 2 and P < 0.05.The differential expressions of lncRNAs were identified by microarray between undifferentiated and differentiated PDLSCs. A total of six samples , which each contained about 1 × 10-ΔΔCt method.For the extraction of total RNA, TRIzol reagent  was used. For quantitative analysis of RNA expression, 1 μg of total RNA was used to synthesize complementary DNA (cDNA) using PrimeScript RT reagent kit (TaKaRa), and the corresponding cDNA was used for quantitative PCR using SYBR green Master MIX (Applied Biosystem) and the ABI 7300 Real-Time PCR System . The PCR program, comprising 1 cycle of 10 min at 95 °C, 40 cycles of 30 s at 95 °C, 30 s at 56 °C, and 1 min at 72 °C, and 1 cycle of 7 min at 72 °C. For the endogenous control, GAPDH was used. The primers were shown in Additional file 5 cells were transferred into 6-well plates, after 24 h, the negative control plasmid or pcDNA3.1-DANCR or pCMV-sh-DANCR were stably transfected into PDLSCs using Lipofectamine 2000 .The pcDNA3.1-DANCR and pCMV-sh-DANCR, as well as negative control plasmid were purchased in GenePharma . And a total of 3 × 102 culture dish, PDLSCs were digested and seeded in 96-well culture plate for another 0, 24, 48, 72 h. Then MTT solution was added and performed as Sun et al. reported [The effect of lncRNA DANCR on cell proliferation was analyzed using an MTT assay. After 24 h of transfection in 100 mmreported .5 cells. After 21 days of induction, ALP activity was measured using the ALP assay kit . Cellular ALP was visualized by using Alkaline Phosphatase Color Development Kit  and the methodology is the same as the manufacturer’s protocol.PDLSCs were separately transferred into 96-well plates, and per well has a density of 1 × 105 cells. After transfection, the cells were then cultured for additional 21 days. We changed the induction medium every 3 days. Finally, the 30 min of alizarin red staining measurement was performed by using Alizarin Red S  at room temperature(24–26 °C). After washing in PBS, the cells were observed using an inverted microscope. To measure the concentration of calcium deposits, the Alizarin Red dye in the PDLSCs was extracted with 400 μl of 10% (w/v) cetylpyridinium chloride in 10 mM sodium phosphate solution for 10 min, and then quantified on a UV-Vis spectrometer at 562 nm.PDLSCs were separately transferred into 24-well plates, and per well has a density of 1 × 10P values < 0.05 were considered as statistically significant.All the statistical analyses were performed by GraphPad Prism (vesion6.0) software. The form of means ± standard deviation (SD) was used for the whole results. Student t-test (unpaired t-test) was used to compare means between differentiated and undifferentiated groups. Differences among the three groups were analyzed via one-way analysis of variance (ANOVA), Tukey’s test was performed in all pairwise comparisons. Additional file 1: Table S1: The primers sequence of qRT–PCR


In Kodaira, Kodama, Kamijo, Kaneko, and Sekijima , an erro