The accumulation of ‘senescent’ cells has long been proposed to act as an ageing mechanism. These cells display a radically altered transcriptome and degenerative phenotype compared with their growing counterparts. Tremendous progress has been made in recent years both in understanding the molecular mechanisms controlling entry into the senescent state and in the direct demonstration that senescent cells act as causal agents of mammalian ageing. The challenges now are to gain a better understanding of how the senescent cell phenotype varies between different individuals and tissues, discover how senescence predisposes to organismal frailty, and develop mechanisms by which the deleterious effects of senescent cells can be ameliorated. Historically, ‘senescent’ cells are the living, but permanently non-dividing, forms of cells that are normally capable of replication within mammalian tissues. Thus, it is possible to have both growing and senescent forms of fibroblasts, keratinocytes, astrocytes, myoblasts, endothelial cells, or vascular smooth muscle cells but not mature neurones, skeletal muscle fibres, or lens fibre cells. Describing these latter cells as ‘senescent’ is usually, but not always, an unfortunate homonym that indicates that the writer is referring to material taken from an aged donor (q.v.).1, although linkages to these appear to be mediated through the mechanistic target of rapamycin (mTOR) axis2. In biological systems where the two processes can be readily separated, principally keratinocytes, senescence can also be shown to be distinct from terminal differentiation3. Senescence limits the capacity for expansion of clones of cells and thus prevents them from accumulating or expanding procarcinogenic mutations, thus acting as anin vivo barrier to carcinogenesis. It is thereby effectively a process-level form of the evolutionary mechanism of action termed ‘antagonistic pleiotropy’. Perhaps unsurprisingly, evidence is gradually accumulating that the short-term presence of senescent cells is beneficial in a range of normal physiological processes, including wound healing and placental development  with3H thymidine or later bromodeoxyuridine. A lack of label incorporation definitively indicated a senescent cell6. In parallel, Hayflick, Smith, Whitney, and their associates used the isolation of individual cells and the measurement of their clonal lifespans to probe the senescent state and yielded impressive data concerning the nature of the molecular machinery8. This non-invasive technique for studying interclonal variation in replicative capacity could fairly be said to have reached its zenith in the work of Shall and colleagues on human and rodent mortal cell populations9.Senescence was famously first reported by Hayflick and Moorhead as a phenomenon limiting the capacity of populations of normal human lung fibroblasts to continue to expandin vivo to show reduced accumulation of senescent cells with calorie restriction10 in the rodent lens; however, this was a technical tour de force). This lack of a utile marker for senescent cells in tissue hampered, and to some extent continues to hamper, the routine detection, quantitation, and localisation of senescent cellsin vivo. The development of the senescence-associated β galactosidase assay by Dimriet al.11 thus represented a significant improvement. Based on a combination of serendipity and classic catalytic histochemistry, it relies on the fact that many senescent cell types have enlarged secondary lysosomes and thus elevated β galactosidase . Because catalytic histochemical techniques were originally developed to produce quantitative data on enzyme activity in tissues (significantly predating the routine use of immunocytochemistry), the senescence-associated β galactosidase assay was readily applicable to fresh tissue sections. Interestingly, alternative techniques based on other aspects of the classic senescent fibroblast phenotype such as the auto-fluorescence associated with the accumulation of lipofuscin are being developed13 and clearly hold considerable promise provided that users do not expect too much .Problematically, these simple techniques did not typically allow researchers to distinguish between senescent and quiescent cells in intact tissue  as markers of senescence. This technique has been used to demonstrate the presence of senescent cells in primate skin and their accumulation with age but remains technically challenging17.In serially passaged cultures of human fibroblasts, senescence primarily occurs because progressive telomere shortening triggers permanent cell cycle arrest through the tumour suppressor gene p53 and the cyclin-dependent kinase inhibitor p21in vitro for many human cell types, including fibroblasts, retinal pigmented epithelial cells, endothelial cells, and urothelial cells20. However, alternative pathways exist based on the action of the cyclin-dependent kinase inhibitor p16 (a selective inhibitor of cyclin D-CDK4/6 kinase pairs)18 or as a direct result of the activation of p53 independent of telomere shortening19. Rodent fibroblastsin vitro enter senescence through a pathway akin to this latter mechanism, which requires the activity of the p19 (p14 in human) ARF tumour suppressor protein  and p16 itself22. Thus, the detection of p16 at the message or protein level can form a useful marker for the presence of senescent cells, although, as with the majority of histochemical techniques, the use of a single marker does not allow definitive assignment of the senescent state23.Telomere-dependent senescence is the primary mode of entry into the senescent state25.This basic network can be linked to a broad series of cellular stressors. These include, but are not limited to, oncogene activation, elevated levels of reactive oxygen species (stress-induced premature senescence), stress signalling in the endoplasmic reticulum (ER stress), glyoxal-induced senescence (metabolic stress), and transforming growth factor beta-induced cellular senescence  is the telomere length-dependent senescence mechanism regularly observed in human cells28. Rather, this appears to be present only in large, long-lived species as an additional tumour suppression mechanism. This trend can be observed within the orderRodentia, in which house mice and rats have long telomeres and active telomerase whereas fibroblasts from the much larger beavers, porcupines, and capybaras have telomeres much closer to human length and lack telomerase activity27.At least two of the three known primary senescence pathways seen in human cells are probably present in mice29. Telomere-associated damage foci similar to those seenin vitro can be observedin vivo as laboratory mice age, suggesting that this mechanism generates senescent cells under normal conditions.However, this does not mean that telomeric effects cannot play causal roles in cellular senescence in laboratory rodents. Although telomerase knockout mice are essentially aphenotypic for several generations because of their long telomeres, Passos and colleagues have shown that damage at the telomere is difficult to repair, resulting in telomerase-positive cells entering a senescent state when they are dosed with gamma radiationin vivo, secondly that they accumulate with age, and finally that an accumulation of senescent cells can have a negative impact. Each is worthy of examination.From its initial discovery, it was postulated that senescence, on some level, was linked with organismal ageing. Modern forms of this hypothesis propose that senescent cells are produced gradually throughout life. These then begin to accumulate in mitotic tissues and act as causal agents of the ageing process through the disruption of tissue function. This conceptual model carries three underlying assumptions: firstly that senescent cells are presentin vitro are assumed to be similarin vivo. Many early reports evaluated findings with the mistaken underlying assumption that cell cultures become senescent because all of the cells divide synchronously for a fixed number of times and then stop. In fact, it has been known since the early 1970s that each time a cell goes through the cell cycle (i) it has a finite chance of entering the senescent state and (ii) this chance increases with each subsequent division30. Thus, senescent cells appear early on if a cell population is required to divide. Indirect demonstrations that senescent cells occurin vivo, accumulate with ageing, and do so at reduced rates in organisms where ageing is slowed  were occasionally published from the 1970s onwards31.A steady production of senescent cells is quite plausible if the kinetics whereby populations of normal cells become senescentHowever, they were technically difficult to perform and correspondingly hard to interpret.33. A central component of this shift is the secretion of biologically active proteins  that have potent autocrine and paracrine activities, a process termed the senescence-associated secretory phenotype (SASP)35. This results in cells that overproduce a wide variety of pro-inflammatory cytokines, typically through the induction of nuclear factor kappa B (NF-κB)36 and matrix-degrading proteins such as collagenase37. Other radical phenotypic changes, such as calcification, have also been shown to occur in some cell types with the onset of replicative senescence38.The presence of senescent cells would be less of a physiological handicap if they behaved in ways indistinguishable from their growth-competent counterparts. Unfortunately, senescence triggers changes in gene expression on (roughly) the same scale as differentiation from one cell type into another41. Such studies demonstrated that senescent cells could, at least potentially, produce significant and diverse degenerative pathology.The individual components of the SASP vary from tissue to tissue and, within a given cell type, can differ depending upon the stimulus used to induce senescence must produce pathology, and a historic weakness of the cell senescence literature was that thein vivo studies essential to testing the causal relationship between ageing and cellular senescence (induced by any mechanism) were lacking. However, the production of transgenic mouse models in which it was possible to eliminate senescent cells has finally made such tests experimentally feasible. Initial studies42 demonstrated first that senescent cells appeared to play a causal role in a variety of age-associated pathologies in theBubR1 mutant mouse and subsequently that either life-long removal of senescent cells or their clearance late in life significantly attenuated the development of such pathologies in these progeroid animals43. This clearly demonstrated that senescent cells can have significant, deleterious effectsin vivo. Interestingly, the removal of senescent cells in this system was not associated with increased lifespan (an observation that demonstrated that it is possible to achieve classic ‘compression of morbidity’ by deleting senescent cells). However, on more conventional genetic backgrounds, attenuated age-related organ deterioration was accompanied by increases in lifespan of the order of 25%44. A justifiable claim can be made to consider these studies ‘landmarks’ in the field in that (i) they demonstrate a causal relationship between senescent cells and ‘ageing’ and (ii) the same mechanism can cause changes associated with ‘ageing’ as well as those associated with ‘age-related disease’. These results have unusually profound philosophical implications for a scientific paper and challenge a fundamental ontological distinction that has been drawn for almost two thousand years between ‘natural’ ageing and ‘unnatural’ disease45.However, the observation that something can produce pathology does not mean that it46.The interaction between senescence and the age-related loss or functional alteration of terminally differentiated post-mitotic cell populations such as skeletal muscle fibres or neurons has been relatively little studied. Such interactions could occur in at least two distinct ways: (i) paracrine effects on neurons or muscle fibres by senescent cells from adjacent mitotic compartments  and (ii) cell-intrinsic changes in phenotype in muscle or neurons resulting from activation of the same pathways that give senescent cells their distinctive phenotype . Given that approximately 30 to 40% of the human body is muscle, either of these could be physiologically highly significantin vivo induces a p21waf-mediated elevation in interleukin-6 production, conversion of euchromatin into heterochromatin, and elevated levels of lysosomal enzyme activity . Such damaged neurons accumulate with advancing agein vivo and that accumulation can be partially rescued by dietary restriction47. The researchers who reported these observations termed the neuronal phenotype a ‘senescence-like state’, which is perfectly accurate but potentially confusing for those with only a cursory interest in the field. A suitably colourless neologism  for such phenotypic changes occurring in cell types that are already post-mitotic would seem to be called for, as would further investigation of their effects on tissue function.In the first case, there is ample potential for senescent cells to trigger inflammatory responses in neuromuscular tissue, which then can be compounded by muscle and neuronal cells, leading to more widespread tissue dysfunction. In the second, it is already known that DNA damage in mature neurons50, they are central players both in age-related mortality and in various forms of morbidity . However, data on their phenotype and behaviour fall far short of what is required if therapeutic progress is to be made in a reasonable timescale. For example, systematic studies on the frequency and distribution of senescent cells in human tissue analogous to those carried out on baboon skin remain to be performed . The result is a literature onin vivo senescence in human tissues that is currently fragmentary and frustratingly difficult to interpret. Even such an apparently straightforward question such as whether the frequency of senescent cells is elevated in the skin of patients with accelerated ageing disorders such as Werner’s syndrome remains unknown. This is not because those in the field do not recognise the utility of such work; rather, it is that studies of this type are considered—in the words of one anonymous but unsympathetic referee—“boring descriptive work”. Similar issues of problem choice have occurred in other branches of ageing research at different times . Thus, linkages to the fraction of senescent cells required to produce degenerative effects in any given human tissue remain speculative and largely based onin vitro data; a figure of about 10% of the total cell population would seem to be sufficient54.Although senescent cells play an important role in tumour suppression and some unsuspected aspects of normal physiologyWhether an enhanced emphasis on basic human studies is a useful parallel-track approach to the pioneering work now taking place in rodent models or an essential next step is a matter of perspective. Many fundamental mechanisms of ageing are conserved between species (such as the insulin–insulin-like growth factor 1 axis), but there are often important species-specific differences. Those inclined to stress the cross-species similarities will be inclined to deprioritise human studies and vice versa.55, it would be surprising if important intra-individual variation did not exist within the general population as well as ‘outliers’ . Although data on differential SASP profiles in response to a senescence stimulus are beginning to enter the literature, they remain fragmentary concerning the senescent cell phenotype in different tissues.Regardless of the species of origin, the extent of variation in the phenotype of senescent cells derived from the same tissue in different individuals is not well characterised. Whilst there are clearly commonalities to aspects of the senescent phenotype such as the SASP57. Recently, navitoclax was shown to provide therapeutic benefit in rodent models of atherosclerosis, mimicking the effects of senescent cell deletion models and reducing the size, number, and overall burden of atherosclerotic plaques58.Despite these gaps, progress is being made towards the development of ‘senolytic’ drugs that can destroy senescent cells—with the goal of duplicating the effects of the transgenic mouse models first in normal animals and eventually in human patients. Initial results seem promising; for example, the BCl2 inhibitor navitoclax was shown to be senolytic in some types of senescent cells  but not others 59. Quercetin also inhibits matrix metalloproteinase activity, another canonical senescence marker60. This demonstrates the potential of an alternative route to dealing with the degenerative effects of senescent cells, blocking their effects rather than killing them, although improved pharmacological profiles are required before such treatments can progress further. The polyphenolic resveratrol also inhibits matrix-degrading enzymes and shows significant anti-inflammatory effects63. Though apparently not senolytic, resveratrol blocks both the SASP in fibroblasts and the pro-calcificatory phenotype in vascular smooth muscle cells64. In addition, it has a range of beneficial effects on post-mitotic cell types, improving muscle mass, strength, and fatigue in rodent models67 and showing beneficial effects in a range of neuronal models68.Dual treatment with the senolytic compounds dasatinib and quercetin mimics the results seen in transgenic mouse senescence clearance models and improves both vasomotor function and vascular stiffness in aged and atherosclerotic mice, although the mechanistic spectrum of action of these compounds is broad and caution needs to be applied in ascribing the improvements in health status seen in senolysis alone62, suggesting that blockade of this pathway may be of general benefit in treating senescence. The potential utility of selective NF-κB inhibition was recently demonstrated inXPF-ERCC1 mutant mice, which show a wide range of progeroid features. In this model, either genetic depletion of one allele of the p65 subunit of NF-κB or treatment with a peptide inhibitor of the NF-κB activator IKK  significantly delayed the onset of symptoms69. Such studies could be usefully combined with other models of progeroid pathology, such asBubR1. Drugs already in clinical use, such as rapamycin and metformin, also modulate the SASP, suggesting that at least some of their beneficial effects are mediated via this route71.Some of resveratrol’s beneficial effects appear to occur through the suppression of NF-κB activationin vitro, suggesting that this shift in the intact organism either results from the accumulation of senescent cells or is a parallel process affecting both senescent cells and those that have been called on to cycle repeatedly72. We have extensive data showing that resveralogues reverse these changes in splicing factor expressionin vitro and rescue multiple features of senescence independently of cell cycle traverse, SIRT1 activation, or SASP modulation. Understanding the mechanism of action of these compounds in greater detail will allow the development of small-molecule anti-degenerative treatments and provide another novel means of improving healthspan, the central goal of modern geroscience.Many aspects of the biology of senescent cells and the relationship of that biology to the aged phenotype remain to be uncovered. For example, RNA splicing factor expression alters profoundly during the ageing of both humans and rodents. This has the potential to compromise the ability of an aged tissue to produce a full spectrum of proteins in response to a physiological challenge (such as cold or heat stress). Changes to alternative splicing are also associated with replicative senescence


Drosophila are offering complementary insights into how these fascinating cells integrate a multitude of guidance cues to fulfill their many tasks and migrate to distant sites to either direct developmental patterning or raise an inflammatory response.Macrophages are multifunctional innate immune cells that seed all tissues within the body and play disparate roles throughout development and in adult tissues, both in health and disease. Their complex developmental origins and many of their functions are being deciphered in mammalian tissues, but opportunities for live imaging and the genetic tractability of  Drosophila macrophages in the contexts of development, phagocytosis, patterning and immune response, and disease states. They discuss the advances allowed by live imaging and fly genetics, and how insights from both insect and mammalian systems can be combined.Wood and Martin discuss the similarities between mouse and  Drosophila embryo, larvae, and pupae are offering useful additional insight into molecular mechanisms, particularly those underpinning how macrophages migrate within tissues and how they integrate several incoming cues to determine their responsive behavior in various circumstances. In this review we briefly describe what is known about the origins of mammalian macrophages and their functions in both developmental patterning of the embryo and during tissue repair, where it seems that embryonic morphogenesis is recapitulated to help rebuild damaged tissues. As some aspects of macrophage function and signaling are not yet tractable in mammals, here we describe Drosophila studies that might help fill the gaps and guide the way forward.Macrophage is a term first coined by Metchnikoff in the late 1800s to describe “big eating” cells that he observed in starfish embryos as they exhibited a foreign body response after he poked the embryos with a rose thorn . MacrophIn the last 10 or so years, various tracking and lineage fate mapping studies in mice have made large inroads into discovering from where all the macrophage-like cells in various tissues are derived. GATA1/2 and PU.1 are key hematopoietic transcription factors that directly interact to repress alternative lineage programs and when PU.1 activity dominates, monocytes/macrophages develop . In largDrosophila and mammals -derived tissue macrophages , which, in turn, is pivotal for equal dispersal of macrophages beneath the embryonic epidermis . A recenDrosophila blood cell: macrophages (plasmatocytes), crystal cells, and lamellocytes. These macrophages can be considered the fly equivalent of vertebrate bone marrow-derived macrophages , and their clearance, was initially missed because both the death and clearance events are relatively rapid (and so appear rare), by comparison with the period over which this neural remodeling occurs . More im corpses . In both corpses ; this ki corpses .Drosophila macrophages function as professional phagocytes within the embryo, efficiently engulfing and degrading large numbers of apoptotic corpses generated during normal development. How macrophages detect, engulf, and degrade apoptotic corpses is an intensely studied field, and once again there appears to be strong conservation of molecular mechanism from the fly to vertebrates. Drosophila macrophages use a battery of receptors including, among others, croquemort, a homolog of the vertebrate CD36 scavenger receptor , play an important sentinel role in the immune system to protect the individual against invading pathogens. Studies in the fly using larval macrophages ex vivo to interrogate the phagocytic machinery required for internalization of bacteria revealed the Nimrod family of receptors, Eater  and NimClt flies , and a rlt flies . Studies. aureus , dSR-C1 (E. coli .Erwinia carotovora (Ecc15), macrophages are required for the induction of the expression of the antimicrobial peptide, Diptericin, in the fat body  in macrophages, which then activates JAK/STAT signaling in the fly equivalent of the liver, the fat body . Followifat body , and macfat body . Expressfat body , and antfat body .Drosophila larva infestation by parasitoid wasps, such as Leptopilina boulardi, has been studied extensively and provides a fascinating model for studying macrophage immune behavior in vivo. Upon parasitization, macrophages rapidly mobilize and differentiate into a specialist cell known as a lamellocyte, which forms a multilayer capsule around the parasitic wasp egg in cooperation with macrophages and a third blood cell type, crystal cells  at the wound  ortholog, Eiger  such as ATP and H2O2, suggesting that these signals can only be permissive factors and that the true attractant is a larger molecule (One fascinating aspect of macrophage biology that can perhaps be best studied in the fly is that of signal integration and prioritization. For a macrophage to efficiently migrate toward a given target it must have the capacity to detect the end-target attractant along with other intermediate cues en route, while integrating these signals with other potentially distracting ones within its environment and prioritizing appropriately to prevent being pulled in disparate directions. This remarkable capacity for navigation using several cues over relatively large distances has been partially studied in vertebrate leukocytes through elegant in vitro approaches , but theF tracks . Becausemolecule . Anothermolecule .Drosophila, where the phagocytosis of apoptotic cells by macrophages is an essential primer for their subsequent inflammatory response to tissue damage and infection (Another aspect of immune cell signal integration where the fly has recently provided a significant advance in our understanding is the process of innate immune priming or “trained immunity.” Emerging evidence from vertebrate studies has demonstrated that innate immune cells can develop a form of immunological memory, a trait previously associated only with the adaptive system (reviewed in nfection . This stnfection . This meDrosophila have revealed other novel roles for macrophages. For example, a recent study uncovered an important physiological role for macrophages in regulating the fly's response to dietary stress. Flies fed a lipid-rich diet display reduced insulin sensitivity and life-span, and both of these effects are mediated by macrophages (Unpaired 2 and Unpaired 3 (Another study, this time using adult flies, has shown that macrophages become stimulated by neuronal injury and accumulate around degenerating distal axons in the wing , demonstrophages . This abrophages  and wherrophages . Recent rophages . Anotherrophages . Anotherpaired 3 . These sUndoubtedly, not all that we learn from studies of macrophage signaling and function in flies will directly extrapolate to what the mammalian macrophage is doing in health and disease, but studies harnessing the live imaging opportunities in fly embryos, larvae, and now pupae provide a powerful model for the study of many aspects of macrophage biology, from the specification and developmental organization of these multitasking innate immune cells through to their many and varied roles at sites of disease. The powerful genetics of the fly will continue to inform vertebrate studies, and the integration of work in both systems will help provide a global picture of how these important therapeutic target cells function in both development and disease in the complex setting of a living organism.


Cellular senescence is an important protective mechanism against cell proliferation and has critical roles in aging and aging-related disease. Recently, one interesting observation is that the protein abundance is higher in senescent cells than that in young cells. So far, some factors were presented to interpret this observation, such as active protein synthesis linked with autophagy, mTOR, and oxidative stress. Here, applying bioinformatic analysis of microRNA profiles in young cells and aging cells, we revealed that globally senescent cells show lower miRNA abundance than that in young cells, suggesting that the repression of protein synthesis by miRNA in senescent cells could be largely attenuated. This finding provides clues that protein accumulation in cellular senescence could be associated with lower miRNA abundance in aging cells. Cellular senescence is the process of accumulative changes in both cellular structure and function that disrupts metabolism, resulting in deterioration and irreversible cell cycle arrest. Cells undergo senesce normally after a certain times divisions in vitro and can also be induced to senesce by toxins, irradiation, and oncogenes, etc. . CellulaRecently, one amazing observation is that the total amount of proteins in senescent cells increased compared with that in young cells, suggesting that protein synthesis seems to be activated in senescent cells –7. So famiRNAs mainly function as negative gene regulators to repress mRNA translation or degrade mRNAs through binding the 3’UTR of targets . As one Here by an integrative analysis of miRNA profiles in young and aging cells, we revealed that globally senescent cells show lower miRNA abundance than young cells, suggesting that the repression on protein synthesis by miRNAs in senescent cells could be largely released. This finding suggests that lower miRNA abundance in aging cells could be another factor in interpreting the phenomenon of protein accumulation in cellular senescence.http://www.cuilab.cn/), which identified miRNA profiles across more than ten types of human cells for the systems study of miRNA stability. In addition to test the difference between young and old cells, we also obtained the data from the organisms. The miRNA expression profiles in young  and aging macaque brain from the GEO database (GEO dataset: GSE18013) were applied to conduct the analysis. To solid this study, we accessed two more datasets, the miRNA expression data in human aging adipose and the umbilical vein endothelial cells from the GEO database (GEO accession number: GSE64566 and GSE37092). These datasets were summarized in We obtained the miRNA expression profiles of young and replicative senescent human umbilical vein endothelial cells from the GEO database (GEO accession number: GSE37092). We obtained the miRNA expression profiles of young and oncogenic H-Ras V12-induced senescent (RIS) IMR90 cells from the CuiLab  for grid alignment and data extraction of the scanned images. Then, we removed the miRNAs that Foreground-Background intensities are smaller than 30, which were considered to be not reliable. Finally, we normalized the miRNA expression data based on the expression level of U6. It should be noted that because these datasets are from different platforms and labs, they could be normalized using different methods. So they could be not in the same scale. But this does not affect the comparison of the aging and young samples within one dataset. The comparison of miRNA expression levels between aging and young cells is performed using the Wilcoxon test, which is a nonparametric test that compares two paired groups and used to decide whether the two population distributions  are identical.TAM software−16, Wilcoxon test, http://www.cuilab.cn/) from oncogene H-Ras V12-induced senescent (RIS) IMR90 cells. Similar with replicative senescent cells, RIS senescent IMR90 cells also show globally lower miRNAs expression level than the untreated cells  (GEO accession number: GSE37092), we found that senescent HUVECs show globally lower miRNAs expression level than young HUVECs .To solid the above finding, we re-performed the above analysis for two more datasets, GSE64566  and GSE37092 . For the GSE64566 dataset, we defined the samples older than 60 in age as the aging group and the other samples as the young group. As a result, the expression level of miRNAs in the young group is significantly higher than that in the aging group in both epicardial adipose  and subcutaneous adipose . For the human umbilical vein endothelial cell, the young cells have significantly higher miRNA expression level than the aging ones (P<2.2×10As the intrinsic tumor-suppression mechanism, cellular senescence acts as a barrier that represses cell proliferation and the malignancy. Therefore, based on the analysis above, we are wondering the functions of miRNAs upregulated/downregulated in aging cells or tissues. It is known that miRNAs can be oncogenic or tumor suppressive . Using TIn summary, we revealed that global profile of miRNAs expression is decreased during cellular senescence. This indicates that miRNAs synthesis seems to be repressed but not activated during cellular senescence. Considering that the main function of miRNAs are to repress the translation of target mRNAs. This may present a new explanation to the phenomenon of protein accumulation during cellular senescence. That is, the global decreased miRNA level leads to the released repression on mRNAs, thus facilitates to increase the protein abundance. We summarized this hypothesis in However, it should be noted that the aging-associated miRNAs identified by transcriptomics is far from completeness. Therefore, developing bioinformatic methods to predict new aging-associated miRNAs would be important in the future. For doing so, algorithms in predicting miRNA-disease associations could be helpful –17.We revealed that globally the level of miRNAs is decreased during cellular senescence, which provided clues for the phenomenon of protein accumulation in aging cells.


As a result, the recently established field of ‘host manipulation’ has seen rapid expansion over the past few decades with public and scientific interest steadily increasing. However, progress appears to falter when researchers ask how parasites manipulate behaviour, rather than why. A vast majority of the published literature investigating the mechanistic basis underlying behavioural manipulation fails to connect the establishment of the parasite with the reported physiological changes in its host. This has left researchers unable to empirically distinguish/identify adaptive physiological changes enforced by the parasites from pathological side effects of infection, resulting in scientists relying on narratives to explain results, rather than empirical evidence. By contrasting correlative mechanistic evidence for host manipulation against rare cases of causative evidence and drawing from the advanced understanding of physiological systems from other disciplines it is clear we are often skipping over a crucial step in host-manipulation: the production, potential storage, and release of molecules (manipulation factors) that must create the observed physiological changes in hosts if they are adaptive. Identifying these manipulation factors,  Event A and the resulting physiological change in the host as Event B, we can now consider the evidence that connects Event A to Event B in the three cases of host manipulation that follow:Toxoplasma gondii via contact with cat faeces (Event A). Following this, testicular testosterone production increases in the rat leading to hypomethylation of the amygdala (Event B). This is suggested to cause the loss of aversion to cat urine in the infected rats )Host-parasite size disparity may also give insight into when the source becomes active. If host-parasite size disparity is great, we may expect a source that the parasite can store, as higher amounts of manipulation factor may be needed to impact host physiology. Alternatively, in host-parasite systems with little size disparity, such as hairworm-insect systems where the parasite induces its terrestrial host to seek and jump in water, the rate of production of manipulative factors may be high enough not to require storage. However, regardless of size disparity, sudden or rapid behavioural changes in the host may necessitate storage of manipulation factors by the parasite, whereas gradual behavioural changes in hosts may only require sustained production. Determining the probability that a parasite’s source of manipulation factor requires storage is important for two reasons: it informs histological searches for the source, and suggests when in the parasite’s life-cycle the source might appear.Finally, it is also important to consider that parasites may repurpose or expand functionality of existing organelles/tissue for generating and secreting manipulation factors. Therefore, histological analysis should focus on existing structures/organelles as well as identification of new structures. Particular interest should be paid to existing organelles already capable of storage in high-disparity, rapid behavioural change host-parasite systems.Earlier studies often use the term ‘proximate mechanism’ when reporting physiological changes in infected hosts. Proximate is an incorrect epithet for these mechanisms. If adaptive behavioural manipulation is occurring in these cases, the physiological changes in the host are actually several steps downstream from the true cause; the manipulation factors themselves and their source are thus likely the true proximate mechanisms. Adopting this alternative perspective provides a bottom-up approach that has the potential to empirically differentiate an adaptive physiological manipulation from other possible consequences of infection. When implemented, this approach will likely accelerate progress in the field of behavioural manipulation.


A classical view of neural computation is that it can be characterized in terms of convergence to fixed-point-type attractor states  to a biophysically detailed level of neuronal systems description .Metastability and its possible functional role both within and outside of behavioral task contexts is further addressed in four specific modeling approaches situated at different spatial scales, ranging from macro/mesoscopic levels . Following up on this theory, Mazzucato et al. show that variation in neuronal ensemble activity may be confined to small subspaces of the whole state space spanned by all the individual units' firing-rates. Moreover, the dimensionality of these subspaces is smaller during stimulus-evoked activity than in the absence of a task. This is in line with empirical studies which report the reduction of neuronal variability upon stimulus presentation  scales, using a clustered spiking model (in which connectivity patterns are heterogeneous) which exhibits metastable states, Tošić et al. proposed a new data analysis technique to identify metastability empirically, which was used to infer metastable states in local field potentials evoked by visual stimuli in anesthetised ferrets.Metastability was also addressed in four studies which provide novel analytical tools and empirical evidence. visual scan paths (Wilkinson and Metta) reveal complex dynamics which possibly reflects underlying metastable neural activity. In Wilkinson and Metta, the authors proposed a theoretical framework, termed the singularity hypothesis, which relies on transient spiral waves which govern persistent neural activity states underlying oculomotor postural control. In general, motor control strategies may be represented in neuronal activity patterns as a complex, distributed spatiotemporal code, which may not be revealed by looking just at neuronal firing rates within recorded ensembles. This is shown in Mao et al. who study behaving rats performing a directional choice task, using a nonlinear decoder to demonstrate how spatiotemporal activity patterns in motor areas increasingly discriminate the animal's choices as learning progresses.Interestingly, Liu et al.; Yada et al.) which further illustrate the rich diversity in methodological approaches to the empirical identification of metastable and transient dynamics. Specifically, Yada et al. propose that repeating spatiotemporal patterns emerging during synchronized bursts of activity in vitro may have their origin in specific sub-populations that become sequentially active in a reproducible temporal order. This result suggests an orchestrated activation of such ensembles that depends on the global state of the network, consistent with spontaneous transitions between metastable states.Finally, spatiotemporal patterns generated by synchronized spontaneous activity in the idle brain are analyzed at different spatial scales in two studies , one of the most intriguing topics currently in systems neuroscience (Moreno-Bote, In summary, this book provides a comprehensive collection of current modeling and data analysis approaches related to the metastable behavior of cortical ensembles. These studies showcase recent efforts for designing a fundamental framework that encompasses the multiple facets of metastability in neural responses, beyond the original use of the concept in the context of statistical mechanics. To conclude, the last chapter of the book reflects on this plethora of approaches and connects them with the question of the functional role of experimentally observed trial-to-trial variability (All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The growing occurrence of bone disorders and the increase in aging population have resulted in the need for more effective therapies to meet this request. Bone tissue engineering strategies, by combining biomaterials, cells, and signaling factors, are seen as alternatives to conventional bone grafts for repairing or rebuilding bone defects. Indeed, skeletal tissue engineering has not yet achieved full translation into clinical practice because of several challenges. Bone biofabrication by additive manufacturing techniques may represent a possible solution, with its intrinsic capability for accuracy, reproducibility, and customization of scaffolds as well as cell and signaling molecule delivery. This review examines the existing research in bone biofabrication and the appropriate cells and factors selection for successful bone regeneration as well as limitations affecting these approaches. Challenges that need to be tackled with the highest priority are the obtainment of appropriate vascularized scaffolds with an accurate spatiotemporal biochemical and mechanical stimuli release, in order to improve osseointegration as well as osteogenesis. Bone is composed of bone tissue and bone marrow encased within the periosteum, a thin strip of soft tissue that envelops the midshafts of long bones, extending to their proximal and distal metaphyses and adjacent epiphyses  this capability can however fail, resulting in permanent defects that can lead to a loss of function. Bone regenerative ability declines with age; therefore, there is a need for In bone defect treatments, the “gold standard” remains bone grafting  into bone tissue engineering strategies has been a crucial progress. The most frequently used are bone marrow stromal cells (BM-MSCs)  and oral cavity MSCs. ASCs have a documented Another attractive stem cell reservoir that could meet bone tissue engineering criteria is the skin basal layer  stimuli as well as in cocultures , secreted locally in the areas undergoing bone remodeling or at the injury sites, cause the migration of inflammatory and precursor cells and/or the activation of osteoblasts and osteoclasts  printing methods can create bioactive or bioinstructive scaffolds, incorporating growth factors or drugs with a spatiotemporal distribution. For instance, the spatial patterns of bone morphogenetic protein (BMP)-2, generated on a fibrin surface using an inkjet bioprinter, was able to differently affect murine muscle-derived stem cells: cells seeded onto the BMP-2 pattern undergo osteogenic differentiation as evidenced by alkaline phosphatase activity, while those seeded outside the BMP-2 pattern remain undifferentiated  plays a key role in angiogenesis during bone development. Several studies investigated the consequence of VEGF, or of a combination of growth factors, on angiogenesis in engineered bone constructs. The co-immobilization of VEGF and angiopoietin-1 on 3D porous collagen scaffolds increased endothelial cell proliferation Rapid prototyping or solid freeform fabrication  was developed in the mid-1980s, providing a high level of control of the construct architecture, flexibility to scale-up fabrication, and assuring reproducibility and standardization of the manufacturing process. Scaffolds developed using conventional fabrication techniques lacks in precision and reproducibility; on the contrary, AMT provides customized scaffolds with precise geometries for replacing damaged or diseased tissues and organs. AMT enables the production of 3D artificial implants using many biomaterials able to meet rigorous performance criteria for clinical use  software or imported from clinical 3D scanners such as X-ray computed tomography, magnetic resonance imaging, and single-photon gamma rays (SPECT). The CAD model is then transformed to a Standard Tessellation Language (STL) file that guides the 3D printer computer system to generate layer-by-layer the object. The manufacturing of a variety of biomaterials fitting the different AMTs allowed the development of scaffolds with tunable properties  fabrication is a peculiar microextrusion technique in which the polymer is distributed through a tool-head installed on an arm or on the Laser-assisted bioprinting involve a pulsed laser source, a receiver substrate for patterning and collecting cells and biomaterials, and a target. An essential element is a laser-absorbing interlayer with a high heat transfer coefficient. Individual cells in suspension are “driven” by directed laser beams and deposited onto a solid surface. This cell-by-cell deposition enables a precise cell micropatterning and improves cell interactions , a family of biopolyesters produced by microorganisms as intracellular carbon and energy storage compounds under unstable growth conditions  can be used to direct guide biological results similar to the traditional approaches that involves chemistry or biomolecules  is cell viability: transplanted cells must survive for a sufficient period in order to perform their biological function. In biofabrication approaches, the printed cells need first to survive during all processing and printing steps. Extreme environmental and culture conditions, changes before, during, and/or following printing can adversely affect cell homeostasis or result in cell damage and death (Leach and Mooney, via solution electrospinning techniques and scaffolds fabricated via 3D printing (Visser et al., Cell viability can be also affected by the biomaterial compatibility. Many hydrogels, which are attractive given their high cytocompatibility, have been applied in tissue engineering strategies (Bryant et al., Bone is a metabolically active tissue with an internal vasculature and osteocytes located no more than 100 μm from an intact capillary (Muschler et al., in vitro growth of endothelial cell on PLGA scaffolds was enhanced on 5–20 μm pore sizes, with lower interpore distance. However, this was in contrast to some in vivo studies that showed as a higher porosity permits for faster bone ingrowth and vascularization. Improved bone formation was observed in hydroxyapatite scaffolds with 300–400 μm pore sizes implanted in rats, suggesting that a fast scaffold vascularization determines an osteogenic microenvironment (Klenke et al., in vitro researches in predicting in vivo results as well as the requirement to assess the best pore sizes for each cell type used for bone tissue engineering. Moreover, porous or rough materials integrate in a less fibrotic, better-vascularized way in comparison with smooth, compact forms.Scaffold pores are essential for the formation of bone tissue, since they enable cell migration and ingrowth as well as nutrient diffusion for cell survival (Karageorgiou and Kaplan, An important reason in implant failure is associated with a foreign body reaction: biocompatible biomaterials become encapsulated and are phagocytized by macrophages. Indeed, it has also been shown that porous biomaterials with pore of 30–40 μm in size displayed excellent healing with a pro-healing functionality of macrophages, regardless of polymer composition or implant site (Osathanon et al., A basic problem for a positive translation of any cell therapy for regenerative medicine purposes is still their large-scale production. In this respect, the bioprocessing phases of the producing process must be reproducible and scalable, in accordance with good clinical and manufacturing practice standards, safe for patients, and economically sustainable (Martin et al., The standard file format used to control AMT bioprinters is STL. While it works for solid objects with small complexity, STL is an unfeasible format if internal pore architecture is an integral part of the computer-aided design (Melchels et al., In vivo animal models can produce several relevant data on bone repair processes and testing the effectiveness of bioprinted or biofabricated bone constructs. These researches generally encompass big numbers of animals that raise ethical concerns and are expensive. The development of ex vivo organ cultures (Smith et al., in vitro and in vivo characterization of bioprinted and biofabricated bone constructs. The improve accuracy of in silico predictive models would also reduce the numbers of animals used in in vivo studies (Fedorovich et al., Modern microscopy techniques have helped in the evolution of biomaterials, from their composition to their interactions (Vielreicher et al., When it is necessary for imaging cells within scaffolds, the limited penetration depths for most microscopy techniques is a major limitation. The introduction of multi-photon microscopy as well as advances in microcomputed tomography has allowed for slice-wise optical sectioning and 3D reconstruction of these constructs. These improvements overcome some of the limitations associated with traditional light microscopy. However, the computing power and storage space necessary to perform, analyze, and collect the data obtained with these methods is massive, even by today’s standards in information technology.in vivo, opening a breakthrough for the clinical translation of this technology, even if to achieve this final goal, further steps and challenges remain (Malda and Groll, Finally, biofabrication is a quickly developing field with the capacity to change drastically the actual medical treatments. However, this idea requires integrated bioprinting platforms able to manage multiple materials to tissue constructs with structural integrity and of clinically relevant scales. Recently, Kang et al.  developeBiofabrication using AMT offers a defined and organized approach for bone tissue generation in comparison with traditional techniques. Nevertheless, there are still significant challenges with biofabrication for the development of clinically relevant bone constructs. These problems are not only relative to the existing limitations in AMT but are also due to the possibility to obtain an appropriate vascularization of the structures as well as correct spatiotemporal biochemical and mechanical stimuli, to maximize osteogenesis and osseointegration. Advances in AMT, computational modeling, medical imaging and microscopy technology, bioreactor design, and biomaterial and drug development are therefore mandatory in order to face difficulties and improve the cost-effectiveness of biofabricated bone for clinical therapy, which at present is poor. Moreover, the correct cell selection for an effective clinical result must be taken into consideration.It must also be underlined that all improvement in bone biofabrication will surely aid the knowledge and understanding of skeletal stem cell biology, cell interactions, and responses to external stimuli for bone development, formation, and remodeling related to the aging of population.MB conceived and critically revised the whole manuscript. MO drafted and revised the section relative to cell sources and biomolecules. MF drafted and revised the section relative to the limits of biofabrication techniques. RP critically revised the whole manuscript.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.


Epigenetic modifications, and particularly DNA methylation, have been studied in many tissues, both healthy and diseased, and across numerous developmental stages. The placenta is the only organ that has a transient life of 9 months and undergoes rapid growth and dynamic structural and functional changes across gestation. Additionally, the placenta is unique because although developing within the mother, its genome is identical to that of the foetus. Given these distinctive characteristics, it is not surprising that the epigenetic landscape affecting placental gene expression may be different to that in other healthy tissues. However, the role of epigenetic modifications, and particularly DNA methylation, in placental development remains largely unknown. Of particular interest is the fact that the placenta is the most hypomethylated human tissue and is characterized by the presence of large partially methylated domains (PMDs) containing silenced genes. Moreover, how and why the placenta is hypomethylated and what role DNA methylation plays in regulating placental gene expression across gestation are poorly understood. We review genome-wide DNA methylation studies in the human placenta and highlight that the different cell types that make up the placenta have very different DNA methylation profiles. Summarizing studies on DNA methylation in the placenta and its relationship with pregnancy complications are difficult due to the limited number of studies available for comparison. To understand the key steps in placental development and hence what may be perturbed in pregnancy complications requires large-scale genome-wide DNA methylation studies coupled with transcriptome analyses. Besides mediating maternal–foetal exchange throughout gestation, the placenta plays a major role in orchestrating maternal adaptation to pregnancy by secreting a variety of steroid and peptide hormones. These placental hormones stimulate maternal physiological changes that are essential for pregnancy success. The placenta is unique in several ways. First, although the placenta is a shared organ between mother and foetus, it is an extra-embryonic tissue and is therefore primarily regulated by the foetal genome. Secondly, the placenta separates from mother and foetus after birth, making it a truly transient organ. For these reasons, the epigenetic mechanisms involved in placental development and regulation of gene expression within this tissue may not be subject to the same lifetime epigenetic constraints as other organs that must function throughout an individual’s life.In humans, from implantation of the blastocyst, the placenta invades the decidua colonizing and transforming the uterine spiral arterioles to sequester a maternal blood supply for efficient maternal–foetal exchange . Invadin3) to cytosine bases, is a process catalyzed by DNA methyltransferases . DNMT1 maintains and repairs established DNA methylation, whereas DNMT3A and DNMT3B are involved in de novo DNA methylation  and the inner cell mass (extra-embryonic mesoderm and endoderm progenitors comprising the villous stroma and blood vessels). The villous syncytiotrophoblast is the major cell type in the placenta and is derived from the trophectoderm via the villous cytotrophoblasts which fuse to form the syncytium (Hypomethylation within the placenta is not uniform but occurs in large domains (>100kb) called partially methylated domains (PMDs) which are regions of reduced DNA methylation that cover approximately 40% of the placental genome =0.432; mean =0.389, P-value=9.552e-08)  (552e-08) . When ona).As previously mentioned, the placenta is composed of several different cell types a.Besides 5-methylcytosine, there are other derivatives, such as 5-hydroxymethylcytosine, that arise from the oxidation of 5-methylcytosine in the process of DNA demethylation . In a reb, Novakovic b) used Illumina arrays to show that DNA methylation increases from first trimester to third trimester with levels of DNA methylation very similar between second and third trimester. The authors suggest that the change in cell composition and differentiation of cells that occurs as gestation progresses may contribute to these differences, as well as the bias of this methodology with respect to the actual methylation sites assessed by the array probes (Novakovicb).One of the first studies to characterize DNA methylation across gestation used HPLC to show that global DNA methylation levels increase with gestational age , intrauterine growth restriction (IUGR), preterm birth (PTB) and gestational diabetes mellitus (GDM) has largely been performed using Illumina Infinium Human Methylation BeadChip Arrays . These sDAPK3, PAPPA2) in common between the two studies that were compared  Senior Research Fellowship (GNT1020749) and NHMRC project grant (GNT1059120) awarded to CTR and TB-M. SB is supported by a NHMRC and Australian Research Council (ARC) Dementia Research Development Fellowship (GNT1111206).


Oncogene addiction is a phenomenon whereby suppression of a driver oncogene is associated with dramatic tumor regression that has been observed in experimental models and in response to targeted therapies . HoweverTo understand our recent results, we must first note that apoptosis is regulated by both pro-apoptotic and anti-apoptotic proteins. The pro-apoptotic protein, BIM (BCL2L11), is a BCL2 family member with three major isoforms  that can be generated from mRNA alternative splicing . BIM worWe found in multiple transgenic mouse models of oncogene  induced acute lymphocytic leukemia (ALL), that BIM was the key mediator of apoptosis observed upon oncogene inactivation . In all Importantly, our results are consistent with other reports that suggest that BIM is the key mediator of apoptosis induced by inactivation of the driver oncogene . For exaTargeted therapy aimed at single oncogenes has not been successful for the treatment for most cancers. One potential solution is to further maximize the tumor cell killing by changing the pro-apoptotic versus anti-apoptotic balance. If BIM is the universal mediator of apoptosis upon oncogene inactivation, upregulation of BIM activity should synergize with targeted agents. In particular, BH3 mimetics, such as ABT-263 and obatoclax, can effectively activate BIM and tilt the balance towards apoptosis. Although the efficacy of BH3 mimetics is modest as single agents, they may potentiate the effects of targeted therapies as part of a rational combination. Thus, a combination of these two different classes of drugs should be tested on a broad scale and hold great potential for the treatment of many cancer types that exhibit oncogene addiction.


Cadmium is a highly poisonous metal and a human carcinogen, but the molecular mechanisms underlying its cellular toxicity are not fully understood. Recent findings in yeast cells indicate that cadmium exerts its deleterious effects by inducing widespread misfolding and aggregation of nascent proteins. Here, we discuss this novel mode of toxic heavy metal action and propose a mechanism by which molecular chaperones may reduce the damaging effects of heavy metal ions on protein structures. Human exposure to toxic metals and metalloids is increasing in many parts of the world due to geochemical contamination and industrial pollution. The heavy metal cadmium is considered genotoxic and neurotoxic, and exposure is associated with lung and kidney cancers. In addition, cadmium may be implicated in the progression of neurological disorders caused by protein misfolding   , assist the folding of proteins into their functional conformation (foldases), or help misfolded and aggregated proteins to regain their native structures (unfoldases) (Finka et al. Although chaperones and other heat-shock proteins are induced by cadmium and other heavy metals or metalloids, it is not clear to what extent the rescuing of proteins contributes to tolerance (Tamás et al. We have previously shown in vitro that during mild heat-denaturation, trace amounts of an aggregation-prone protein acted as proteinaceous infectious seeds that could dramatically accelerate the misfolding of another less thermolabile protein in excess. Remarkably, the presence of an active unfolding/disaggregating chaperone system, that effectively neutralized the seed formation, could optimally prevent seed-induced aggregation of the less labile abundant protein species (Ben-Zvi and Goloubinoff Our recent studies in yeast cells have demonstrated that cadmium (Jacobson et al.


Use of electronic clinical trial portals has increased in recent years to assist with sponsor-investigator communication, safety reporting, and clinical trial management. Electronic portals can help reduce time and costs associated with processing paperwork and add security measures; however, there is a lack of information on clinical trial investigative staff’s perceived challenges and benefits of using portals.The Clinical Trials Transformation Initiative (CTTI) sought to (1) identify challenges to investigator receipt and management of investigational new drug (IND) safety reports at oncologic investigative sites and coordinating centers and (2) facilitate adoption of best practices for communicating and managing IND safety reports using electronic portals.CTTI, a public-private partnership to improve the conduct of clinical trials, distributed surveys and conducted interviews in an opinion-gathering effort to record investigator and research staff views on electronic portals in the context of the new safety reporting requirements described in the US Food and Drug Administration’s final rule . The project focused on receipt, management, and review of safety reports as opposed to the reporting of adverse events.The top challenge investigators and staff identified in using individual sponsor portals was remembering several complex individual passwords to access each site. Also, certain tasks are time-consuming  due to slow sites or difficulties associated with particular operating systems or software. To improve user experiences, respondents suggested that portals function independently of browsers and operating systems, have intuitive interfaces with easy navigation, and incorporate additional features that would allow users to filter, search, and batch safety reports.Results indicate that an ideal system for sharing expedited IND safety information is through a central portal used by all sponsors. Until this is feasible, electronic reporting portals should at least have consistent functionality. CTTI has issued recommendations to improve the quality and use of electronic portals. Safety reporting is a necessary element of clinical trials to help ensure patient safety during the investigation of a new drug or medical device. With advances in technology, safety reporting, along with other clinical trial data reporting, is moving to electronic formats from being largely paper-based. To encourage electronic submissions and integration of other technological capabilities into trial document management, the US Food and Drug Administration (FDA) has previously released guidance on electronic submissions and source data ,2.Aligned with this progress is the increased use of electronic portals to facilitate communication between sponsors and the investigative staff during clinical trials. Few publications in the scholarly literature have focused on portals; however, it is acknowledged within the clinical research field that use of portals is proliferating. Typically, clinical trial portals are developed by sponsors or contract research organizations (CRO) to provide a centralized location for trial-specific documents and information . Using clinical trial portals can reduce the time and cost associated with processing paperwork, among other advantages. The portal also provides increased security with document management and communication through the use of log-in identification and password protection. Once the portals are developed, investigators are given access to the portal through specific log-in credentials. Investigators can access trial-specific documents provided by the sponsor or CRO. Throughout the trial, investigators periodically log in to review safety reports.The Clinical Trials Transformation Initiative (CTTI) is a public-private partnership whose mission is to develop and drive adoption of practices that will increase the quality and efficiency of clinical trials. CTTI initiated the investigation new drug (IND) Safety Advancement Project to investigate barriers to implementation of the FDA final rule on expedited IND safety reporting  and propose solutions to address identified barriers, from the perspective of both investigators and sponsors. The project focused specifically on oncology research, where these issues tend to be most acute.Evidence gathered during the project indicated that investigators have difficulty using clinical trial portals and that portals may contribute to confusion and burden investigators’ experience related to the IND safety reporting system. Therefore, the CTTI project team sought to identify best practices for managing IND safety reports using electronic portals and formulated recommendations based on data collected through evidence-gathering activities.The IND Safety Advancement Project team included 20 individuals representing a wide range of stakeholders, including industry, academic institutions, institutional review board (IRB), regulatory, patient advocate, and other perspectives; all groups were considered equal partners. The primary focus of the project was to investigate barriers related to the lack of implementation of the final rule (21 CFR 312) and provide recommendations for better compliance; however, a portion of this project was specifically devoted to addressing the use of clinical trial portals. The team employed 3 main research strategies to gather evidence: surveys, expert interviews, and an expert meeting.An online survey was designed to assess challenges related to cancer researcher management of IND safety reporting processes, with a subset of questions about the specific challenges related to use of electronic portals to manage safety reports . The surQuestions on current issues:If IND safety reports are distributed via a sponsor safety reporting portal, do you have difficulty accessing the IND safety reporting portal?Please describe the difficulty you have accessing the IND safety reporting portal.Questions on suggestions for improvement:What things about the current IND safety reporting system should be changed?If you were starting from scratch, what would an ideal IND safety reporting system look like?A total of 20 in-depth, hour-long interviews were conducted by a professional interviewer as an opinion-gathering effort to more fully understand sponsor and investigative staff perspectives on the management of IND safety reporting processes. Interview participants who were considered experts in the topic were approached by CTTI. Survey respondents were also able to volunteer for interview participation. In January and February 2015, 13 individuals representing investigative staff working on oncology clinical trials and 7 pharmacovigilance leaders from 5 large global pharmaceutical companies were interviewed. Prepared questions were included in an interview guide . The goaThe project, including the surveys and interview guides, was designated as exempt research by the Duke University IRB.The IND Safety Advancement Project team analyzed survey results and interviews, developed draft recommendations based on responses, and presented this information at a 2-day expert meeting in July 2015 attended by 47 individuals representing a variety of clinical trial stakeholders. Discussion from the meeting was used by the project team to refine the recommendations through iterative, consensus-driven discussion, and they were approved by CTTI’s Executive Committee prior to official release (December 2015) .The survey had 201 respondents. The majority of the respondent population had academic or community-based research backgrounds with more than 10 years of clinical trial experience. A majority of the respondent-affiliated organizations conducted more than 30 studies concurrently and represented all phases of clinical trial development sponsored by industry and government. Full results of the survey and interviews are being reported concurrently elsewhere . This maResponses from investigators and other study staff indicate that 80% (33/41) of investigators and 92% (133/144) of study staff received IND safety reports through portals. Approximately half  of investigators and 44% (64/144) of staff reported difficulty accessing sponsors’ IND safety reporting portals. When survey respondents were asked to provide free-text responses specifying difficulties encountered accessing a portal, a common answer was the problem of remembering passwords for numerous individual sponsor portals. Other difficulties voiced by respondents were challenges with operating systems, software compatibility, and differing application versions. Respondents indicated that many interfaces are difficult to navigate and do not incorporate intuitive design elements. Additionally, many portals have slow processing times, and their applications often crash or fail. For these reasons, downloading reports can be time-consuming for the investigative staff. Finally, respondents noted that generic email notifications are not particularly helpful and many choose to block these emails. The top-rated challenges were as follows:Remembering numerous, complex passwords for individual sponsor portalsEncountering problems related to different operating systems, software compatibility, and application versioningNavigating through nonintuitive user interfaces and slow sitesEncountering log-in or site issues due to investigative staff turnoverReceiving too many generic email notificationsTime-consuming process of downloading reportsAlthough distribution of streamlined reports via electronic portals was intended to improve the efficiency of the safety reporting process, interviewees from investigative sites reported that they continue to receive an unmanageable volume of IND safety reports. Active sites described the volume of reports as “staggering.”Respondents described a number of benefits related to use of portals for safety reporting, including automatic notifications of trends or unexpected adverse events, which help guide treatment decisions for patients. Investigative staff indicated that it was easier to identify risks when reports were submitted through the portal. Summary reporting and the defined attribution and causality available on the portal can help filter the safety information for study staff. With enhanced signal detection, the investigative staff can identify information that may generate important changes to the protocol or consent and help them make determinations on the utility of the study and other risk/benefit assessments. Additionally, electronic reports are more efficient and easier to retain and track.The attributes displayed in Overall system functionality:Cross-browser compatibility; portal works seamlessly with all commonly used browsersOperating system independentMobile-friendlyQuick report download time Simplified security management system User interface:Intuitive, easy-to-navigate interface requiring few clicks to access safety reports directly or via hyperlink contained in an email notificationFlexibility within the portal for use with varied institutional processesReport management and analysis:Print reports or download multiple reports with one click to a compact disc, computer, or electronic investigator site fileFilter reports by event so follow-up safety reports do not appear as new eventsSearch and display safety reports using custom dates, by country of origin and/or event nameExport single reports as well as aggregated dataDrill down to access single reports and write-upsReports remain visible for the life of the trialReport notification, acknowledgment, and verification:Ability to batch safety report notifications (per day/week) per investigative site user’s preferenceAbility for principal investigator to delegate the task of accessing safety reports via portal to another person at the siteEasy acknowledgement of safety reports by investigative site staff Ability to send and record acknowledgement of a safety report only once across multiple trials for the same investigational product yet still show the report under each trialEnd-to-end audit trail that can be printed and saved or stored for future reference by both the sponsor and investigative siteAbility of the sponsor to document delivery of reports within the portal if an alternative means of reporting is required Two-way communication between the investigative site and sponsor regarding safety reportsAs described elsewhere , investiSurvey results indicated that an ideal system for sharing expedited IND safety information with investigative sites is through a central portal used by all sponsors in order to improve efficiency and reduce paperwork burden . This approach may reduce the number of passwords and avoid technological issues  by standardizing the use of a central portal. While literature describing clinical trial portal use is limited, some reports validate survey respondents’ perceived obstacles, indicating that investigative staff and investigators struggle to recall 7 to 15 passwords per individual user . Until uThe attributes listed in Browser and operating system independence so that all users can access the portal regardless of software preferencesAn intuitive interface that is clearly labeled and provides easy navigationAbility to filter and search reports to quickly access only the relevant documents needed at the timeIncreased functionality to batch reports so that all files can be downloaded simultaneouslyAbility to acknowledge receipt quickly with a check-box option and to update this acknowledgement across multiple trials for the same productPortal features to more accurately track audit trails can be particularly helpful, as reports can be categorized or searched by date, number, compound, trial number, upload/availability date, download/access date, identity of users who accessed the report, and any actions  conducted by a specific user. As indicated in To support appropriate use of the portal, CTTI also recommends that there be improved education for investigative sites including guidance not only on portal functionality but also regarding best practices for incorporating portal management into site report management processes. Finally, we recommend usability testing for portal-related educational material.The main limitation of the study that should be acknowledged is the small sample size that was selected by convenience, which may be susceptible to bias. Additionally, survey response rate was not able to be determined. CTTI acknowledges these shortcomings and that this research is exploratory and qualitative in nature and not statistically measureable. However, change often begins with small steps, and it is our hope that this research sparks a broader discussion across the industry. CTTI encourages additional research on this topic. The perspectives described in this manuscript are descriptive; in order to be conclusive, an appropriately powered study would need to be conducted. CTTI and other independent organizations cannot require that sponsors adopt recommendations nor mandate inclusion or standardization of portal features, which is why we urge portal developers to consider the perspectives of the portal users to drive change.Industry technology companies are investing in integrating sponsor trial portals with enhanced capabilities including interactive voice response, electronic data capture, clinical trial management systems, and investigator databases ,10. The The suggestions provided in this manuscript have been released as CTTI recommendations . CTTI be


Critical Care [We read the editorial entitled “Non-antibiotic treatments for bacterial diseases in an era of progressive antibiotic resistance” by Steven M. Opal published in cal Care . We wantEscherichia coli as recombinant proteins and properties such as rapid killing, specificity, ability to kill antibiotic persisters, low rates of resistance and profound antibiofilm activity [In the editorial, six possible options have been listed as non-antibiotic inhibitors of bacterial growth. We agree that these are viable options which include devices and chemical or biological compounds for addressing the problem of bacterial drug-resistance. Although we understand that this is not an exhaustive list of possible options, we would like to suggest that bacteriophage “lysins”, including both endolysins (which help the phage in releasing the intracellular virus particles by cleaving the peptidoglycan from inside) and ectolysins  can also be considered as alternative options for treating drug-resistant bacteria. When added to a bacterial culture in broth or buffer, the lysins kill bacteria rapidly by cleaving peptidoglycan, a cell wall component specific to bacteria. Because of their specific nature, lysins kill only the target bacteria and thus do not alter the microbiome. Lysins can be easily produced in activity –4 make tactivity –5. Antibactivity . Thus, k I heartily agree with the comments made by Sharma and Paul regarding the potential contribution of phage-medicated endolysin as a novel therapeutic option against multiple antibiotic drug-resistant (MDR) bacterial pathogens. Phage employ a two-component enzyme system, known as holin and endolysin, to allow newly generated phage progeny to exit the dying bacterial cell and invade other susceptible host cells. Holin is a bacterial cell membrane permeablizing enzyme that creates pores within the inner membrane of bacteria, while endolysin destroys the cross-linking of the peptidoglycan layer that constitutes the outer membrane. Holin sets the pace of lysis to correspond to the maximum rate of phage synthesis just before the parasitized bacterial cell dies. At that moment endolysin floods the outer membrane, thereby lysing the cell allowing the new generation of phage particles to be released into the environment .Recombinant forms of lysins have been synthesized and are now being used therapeutically in experimental models of bacterial infection , 7–9. Th


Undergraduate nursing placement in aged care is forecast to grow in importance with the increasing aging population, and to help to reverse trends in student lack of interest in gerontology careers. However, there is a need to better understand undergraduate nursing students’ experiences on placement with older adults, as well as key features of quality learning within residential aged care. The aim of this study was to explore how nursing students understand learning within residential aged care.This qualitative study used a participatory action research approach, and this paper reports on the thematic analysis of data from one cycle of undergraduate nursing placement in a Canadian residential aged care setting, with two groups of 7–8 students and two university instructors. Staff and residents at the research site were also included. Researchers interviewed both groups of students prior to and after placement. Instructors, staff and residents were interviewed post placement.Students commenced placement full of apprehension, and progressed in their learning by taking initiative and through self-directed learning pathways. Engagement with residents was key to student learning on person-centred care and increased understanding of older adults. Students faced challenges to their learning through limited exposure to professional nursing roles and healthcare aide/student relationship issues. By placement end, students had gained unique insights on resident care and began to step into advocacy roles.In learning on placement within residential aged care, students moved from feelings of apprehension to taking on advocacy roles for residents. Better formalizing routes for students to feedback their unique understandings on resident care could ensure their contributions are better integrated and not lost when placements end. HowThis is further complicated by challenges in developing quality clinical learning environments for nursing students to learn and work with an older population. A growing shortage of nurses in Canada , means t"Therefore, RAC is a key setting to kindle student interest in the care of older adults, while developing essential and advanced proficiencies in contemporary nurse practice."Despite its limited use thus far, residential aged care (RAC) is forecast to grow in importance as a site for undergraduate nursing education . PlacemeTo ensure placements in aged care settings contribute to the preparation of an age-friendly nursing workforce, the diverse influences on undergraduate student perceptions of practicum learning in aged care and with older adults must be explored. Evidence already suggests that students tend to find these placements “unsatisfactory and/or unsettling” and experience a sense of powerlessness on placement, reinforced by financial constraints, an unaccommodating learning/working environment, and a lack of readiness to encounter and care for ageing bodies .Clinical placement is a key contributor to cementing student attitudes , and mucThis paper reports on one 13-week undergraduate placement cycle that occurred in the summer of 2016. The study followed a participatory action research (PAR) design, involving cyclic, iterative processes for investigation, action, reflection and collaboration in the inquiry  and are The first stage of this research involved the collaborative and ongoing development of a person-centred placement model aimed at enhancing the interests of undergraduate nursing students in working with older adults. In the spring of 2016, a working group consisting of university educators and researchers, site staff, and residents met in seven 1-h sessions to begin to co-develop the placement model. The working group developed and planned a person-centred placement model for the upcoming student placement, incorporating elements such as the concepts of person-centred care and intergenerational learning; student and resident orientations; communication of student skills and developing capabilities; and roles and expectations of residents, staff, and students.Two groups of undergraduate nursing students, with 1 instructor respectively, were placed at two supportive living facilities owned and operated by the same non-profit provider and co-located on the same campus. The 13-week practicum cycle  occurred in the summer of 2016.At the intervention site, eight students and residents were matched by the working group and structured time was accommodated in the student schedule. Students spent 1.5 days per week at the sites and worked with healthcare aides (HCAs), particularly during morning care routines. Students also worked one-to-one with residents on a care plan assignment.The second site used a traditional clinical placement model. Here, seven students chose which residents to work with and their time with residents was less structured. Like Facility 1, students spent 1.5 days per week at this site, worked with HCAs, and worked one-to-one with residents on a care plan assignment.While there were some variances between the placement models at site 1 and 2 in relation to orientation to site, intentionality and focus in the student–resident interactions, in general the placement model at the intervention site was not fully implemented and there were minimal differences between groups. Therefore findings for both groups are presented together to further understanding of student learning within RAC.n = 15) and instructors (n = 2), and with site staff (n = 8) and residents (n = 5) involved at the intervention site. Instructors in this study were experienced RNs employed by the university, on a contractual basis, to accompany and facilitate undergraduate student learning during clinical placement. Student participants were second year, first term undergraduate students undertaking their first clinically focused placement. Staff included were HCAs, licensed practice nurses (LPNs) and site management .Qualitative data was collected with both groups of students tunity” Comparisons of hands-on nursing versus ‘sitting in the office’ within the data highlight how hands-on care is also seen as contact with residents and tied closely to nursing professional identity. With little exposure to RNs working in the setting and with an understanding that RNs work in more managerial positions, students explained that they wanted to work in future positions that involved contact with residents – and felt that the RN role in RAC would not provide them with that. Aged care nurses themselves feel less satisfaction with their roles, due to greater responsibilities and lack of role clarity, and this has been linked to a limited capacity of some RAC sites to support clinical placement . Care muGreater exposure to positive professional nursing roles could help promote the reality of nursing as a profession involving a broader spectrum of knowledge and services, including administration and case coordination. Abbey et al. (, p. 17–1During placement, students engaged one-to-one, spent extra time generally with residents and formed relationships with older adults living in the aged care setting. These opportunities helped students develop their understandings of person-centred care and gain greater appreciation for older adults. Aud et al.  indicateSpending time with residents facilitated unique insights for students, who used these to step into resident advocacy roles. At both sites, students worked to integrate their unique insights into care delivery through using the extra time they had with residents to build personal relationships, and deliver person-centred care and individually targeted interventions for residents. Feeding back these insights at the organisational level was more problematic. As outsiders, students have little power to enact change in resident care or access formalised processes within the organisation. Elliot et al.  discusseWhere students in our study described being able to build different types of relationships with residents than staff, the potential of student presence on placement supporting person-centred care and resident advocacy is highlighted. Intergenerational service learning initiatives align learning objectives with community-identified priorities  and haveThis study focused on two undergraduate student groups from the same university within the supportive living settings of one RAC organization, therefore the generalizability of the findings is limited. However, the study provides insights on student learning trajectories, integration of staff mentorship to support student learning and the benefits of student-resident engagement opportunities within placements in RAC. There is a need for further investigation into the potential benefits of communicating self-directed learning pathways to students, methods for better integrating professional nursing roles into student learning in RAC and formalizing feedback of unique contributions gained through student-resident engagement opportunities.This study found that taking initiative was important for undergraduate nursing students to move from initial apprehensions to create their own learning opportunities and begin to step into resident advocacy roles while on placement in RAC. Similar student trajectories on placement within RAC are reported in the literature, and methods to enrich mentorship and learning environment are highlighted , 22, 26.Placements in RAC can better support positive learning progression pathways through including RN and LPN job shadowing experiences for students. However, where there are consistently low numbers of senior staff in RAC to contribute to student learning, there must also be real work accomplished on affirming and communicating the value of care aide competencies to students training to be RNs and easing the working of this key relationship within placement. Shaping learning progression pathways within aged care placements as positive with clear guidelines on how sites can contribute to student mentorship could provide greater opportunity for students to gain a more fulsome understanding of the nursing profession within RAC. This may help set the groundwork for students to better picture themselves working in these settings in the future.Including student-resident engagement opportunities on placement was found to help students build their understanding of older adults and learn aspects of person-centred care. Where students had extra time to spend with residents and as these interactions were framed educationally, students described building more personal relationships with residents than staff and gaining unique insights into the lives of residents. Better formalizing and supporting this extra time and attention students spend with residents while on placement could facilitate the unique contributions of students into care delivery.