Cancer has traditionally been considered as a disease resulting from gene mutations. New findings in biology are challenging gene-centered explanations of cancer progression and redirecting them to the non-genetic origins of tumorigenicity. It has become clear that intercellular communication plays a crucial role in cancer progression. Among the most intriguing ways of intercellular communication is that via extracellular vesicles (EVs). EVs are membrane structures released from various types of cells. After separation from the mother membrane, EVs become mobile and may travel from the extracellular space to blood and other body fluids.Recently it has been shown that tumour cells are particularly prone to vesiculation and that tumour-derived EVs can carry proteins, lipids and nucleic acids causative of cancer progression. The uptake of tumour-derived EVs by noncancerous cells can change their normal phenotype to cancerous. The suppression of vesiculation could slow down tumour growth and the spread of metastases. The purpose of this review is to highlight examples of EV-mediated cancer phenotypic transformation in the light of possible therapeutic applications. Cancer has been traditionally viewed as a consequence of multistep mutations of genetic material that result in transformation of normal to malignant cells. However, nowadays the mainstream paradigms of cancer development and progression are shifting from strictly genocentric towards epigenetic and other nongenetic interpretations. It is thus relevant to explore the possibility that a normal cell could become malignant without previous genetic mutation. In this article several mechanisms of phenotypic transformation are presented mainly involving transfer of membrane attached receptors for growth factors, RNA molecules or even lipids 3It was suggested that intercellular communication plays a crucial role in cancer progression.in vivo and in vitro.i.e. urinein vitro and in vivo represent a mixed population of various sizes and origins. To date no consensus regarding their classification and nomenclature was reached to distinguish between different types of vesicles. In this work we do not consider the apoptotic bodies  which are released from the cell in the final stage of programmed cell death.It is considered that EVs are membrane-enclosed compartments, released into the surroundings of practically all cell types, both The content of EVs depends on the cell of origin and the mechanism of vesicle generation. They were found to transfer surface-bound receptors and their ligands, proteins, genetic material, infectious particles, prions and probably even organelles between cells.23Tumour-derived EVs  represen2The exact mechanisms underlying the formation of EVs have not yet been fully elucidated, but it seems that vesiculation can be either an extremely well regulated process, or a random, non-specific event associated with, for example, disintegration of the plasma membrane. It is important to realize that general mechanisms of membrane vesiculation can also play a pivotal role in the progression of disease.Membrane vesiculation takes place in the last phase of membrane budding when the bud is pinched off from the membrane to become a free vesicle . Budding43There is a codependence between membrane shape and structure; moreover, membrane curvature is determined by the shapes of membrane constituents and their interactions.51Additionally EVs seem to be enriched in proteins and lipids associated with membrane rafts.552+ inhibiting translocase, activating scramblase and resulting in loss of membrane asymmetry57Other pathways leading to curvature and subsequent budding of membranes include an increase of intracellular CaMembrane budding can be followed by membrane fission, which is still a subject of some debate, but several ideas supporting the pivotal role of endophilin I and dynamin in this process have been suggested.59EVs can also be formed in processes distinct from those already mentioned. EVs smaller than 100 nm, usually called exosomes, are formed by exocytosis after the assembly of several endosomes into a multivesicular body, exiting the endosomal pathway and fusion with the plasma membrane .50,60,6162It is indicated that EVs interact with the membranes of recipient cells. The precise mechanisms of uptake of EVs are still poorly understood, yet it is becoming increasingly evident that their uptake can induce activation of specific signal transduction cascades and thereby influence the physiological or pathological state of recipient cells.63Several types of interactions were proposed involving adhesion of vesicle molecules to cellular surface receptors (receptor-mediated uptake), endocytosis (phagocytosis) and fusion with the plasma membrane.64et al.Potential receptor candidates for interaction with EV-membranes are, notably, receptors for phosphatidylserine. One such receptor is the T-cell immunoglobulin and mucin-domain-containing molecule (TIM) that was described as mediating vesicle uptake.et al. showed that exosomes preferentially fuse with the membranes of tumour cells and that in these interactions the microenvironmental pH acts as a key factor by modulating the lipid compostition of the cell and exosome membranes , changes in expression of EGFRvIII-regulating genes , morphological transformation and increase in anchorage-independent growth capacity.75Al Nedawi et al., where they detect tumour-specific EGFRvIII in serum EVs of glioblastoma patients.24Similar findings were reported in a study by Skog A mechanism that controls metastatic progression through the EV-mediated transfer of another receptor, tyrosine kinase MET, has recently been described. EVs with oncoprotein MET from highly metastatic melanomas increased the metastatic behaviour of primary tumours by permanently educating and mobilizing bone marrow progenitors.76et al.77Another example of EV-mediated protein delivery in tumour progession has been described by Sidhu Many other proteins have been identified in EVs shed from cancer cells, including among others vascular endothelial growth factor (VEGF)80et al. reported that metastatic gastric cancer cell line releases EVs enriched in let-7-miRNAs, known to negatively regulate Ras genes, leading to maintenance of their oncogenesis.91Recently it has come to light that messenger RNA (mRNA) and various forms of non-coding RNA, such as microRNA (miRNA), act as key players in information transfer between cells.EVs have been found to transfer DNA between cells, but it is important to keep in mind that EV fractions can also consist of apoptotic bodies, known to contain DNA fragments, possibly contributing to genetic changes and tumour progression.93Mitochondrial dysfunction and especially dys-functions caused by mutations of mtDNA have been implicated with a wide range of age-related pathologies, including cancer.95et al. incubated EVs derived from human medulloblastoma cells and enriched in retrotransposon RNAs, especially HERV-K, with HUVEC cells.93A large part of the mammalian genome is derived from ancient transposable elements, such as DNA-transposons and retrotransposons. While DNA-transposons amplify without any RNA intermediate, retrotransposons need reverse transcriptase to retrotranscribe them before integration into the genome.Sphingomyelin is a major membrane phospholipid, mostly localized in the outer leaflet of the mammalian plasma membrane.100et al. showed the importance of sphingomyelin transfer in cancer progression.101Kim e.g. heparin) are suppressors of membrane vesiculation and can therefore have anticoagulant, antimetastatic and anti-inflammatory effect.105It has been shown that heparin, usually used for the treatment of thromboembolismsRecent investigations revealed that invasive tumours can be spread in the body not only by metastases travelling along tissues or being transported by body fluids and so seeding new tumours after anchoring to targeted tissues. Tumours can also be spread by much smaller carriers in the form of EVs containing genetic information or mutant growth factor receptors that are permanently active and provoke over-inducing signalling of cell division. Transfer of such vesicles can occur over short distances to neighbouring cells or long distances by body fluids. By finding appropriate target cells the transferred transforming molecules can induce cell transformation and cause cancer progression most efficiently in already immortalized precancerous or stem cells. As tumor cell transformation is usually a multistep process including several consecutive mutations it can be concluded that the transfer of a transforming molecules can serve as one of the steps in this process. By carrying certain enzymes such as metalloproteinases, the EVs can adapt the microenvironment of tumour cells in favour of metastatic dissemination or implantation into certain tissues. Blocking the spreading of EVs, by the use of molecules attaching the vesicles to the vesiculating cells could possibly slow down tumour growth or the spread of metastases. On the other hand, screening of cancer genetic markers transported by EVs could improve diagnostic methods for detection of certain cancerous diseases. A thorough understanding of the biological mechanisms involved in intercellular communication by EVs could provide a key complement to genetic factors in determination of cancer progression, while their controlled manipulation will likely develop into a powerful weapon in the battlefield of oncology.


Specifically, network synaptic potentiation increases dramatically with high cholinergic concentration and decreases dramatically with low levels of acetylcholine. We demonstrate that this phenomenon is robust across variation of many different network parameters.Although sleep is a fundamental behavior observed in virtually all animal species, its functions remain unclear. One leading proposal, known as the synaptic renormalization hypothesis, suggests that sleep is necessary to counteract a global strengthening of synapses that occurs during wakefulness. Evidence for sleep-dependent synaptic downscaling  has been observed experimentally, but the physiological mechanisms which generate this phenomenon are unknown. In this study, we propose that changes in neuronal membrane excitability induced by acetylcholine may provide a dynamical mechanism for both wake-dependent synaptic upscaling and sleep-dependent downscaling. We show  synaptic renormalization hypothesis, suggests that waking results in a global increase in the strengths of connections in the brain, a phenomenon which is unsustainable because stronger connections consume more energy and take up more space. The function of sleep, according to this hypothesis, is to downscale or “renormalize” connection strengths. While mounting experimental evidence confirms that sleep-dependent synaptic downscaling does occur, we still do not know what biophysical mechanism causes it. In this paper, we show computational results which indicate that the neuromodulator acetylcholine may have a key role to play in sleep-dependent synaptic downscaling. If confirmed experimentally, these findings will help to unravel the mystery of sleep.The function of sleep is one of the greatest mysteries in contemporary neuroscience. Nearly every species of animal requires it, yet we do not know why. One idea, known as the  Sleep is crucial for normal cognitive function as evidenced by the many cognitive impairments associated with chronic sleep loss in vivo studies finding increased slope of evoked LFP/EEG responses after wakefulness and decreased slope following sleep in rats Multiple lines of empirical evidence supporting the synaptic renormalization hypothesis have recently emerged Exactly how synaptic downscaling is induced during sleep is an open question. One suggestion is that the repeated alternation of depolarized “up” states, reflecting the simultaneous activity of many neurons, and hyperpolarized “down” states, reflecting fewer active neurons, observed to occur at approximately 1 Hz during SWA may induce long-term depression (LTD) of synapses A further hypothesis is that differences in the neuromodulators available during waking and NREM sleep states may contribute to the opposing effects of wakefulness and NREM sleep on neuronal potentiation levels In the present study, we build upon previous work to develop a new theory for synaptic downregulation during NREM sleep that highlights a role for differing cortical network dynamics during wake and NREM sleep states. This theory relies upon previous findings showing that acetylcholine (ACh) modulates the phase-dependence of neural responses in cortex Our new theory concerns the effect of increased synchronized network activity during NREM sleep on the strength of synaptic connections. In particular, we posit that although this increase in synchronized network activity strengthens some individual synaptic connections, it weakens others. Further, and critically, this weakening is more pronounced when an animal is experiencing NREM sleep (more synchronized activity) than when an animal is awake (less synchronized activity). Supporting this novel hypothesis, we show that a computational model employing these dynamic, physiologically-plausible mechanisms is fully able to account for synaptic renormalization during NREM sleep.We simulated the effects of ACh on synaptic potentiation in cortical networks consisting of 1000 neurons (20% of which were inhibitory). Each neuron was described by a recently-developed cortical pyramidal cell model and delay their spike firing in response to synaptic input Switching PRCs of synaptically coupled neurons from Type II to Type I has been shown to dramatically affect the synchronization of neuronal networks. Specifically, simulated large-scale neuronal networks whose cells have Type II PRCs have been shown to synchronize much better than neuronal networks composed of cells with Type I PRCs re-wiring probability. Since both maximum synaptic strength and network connectivity structure are known to dramatically influence neuronal network dynamics, we explored a wide range of values for We investigated how the differential effects of ACh on network synchrony influenced overall network synaptic potentiation when synaptic strengths evolved according to a spike-timing dependent plasticity (STDP) rule. In our network simulations, synaptic strength values were initialized to an intermediate value and then allowed to evolve, according to the STDP rule, over the interval High simulated cholinergic modulation switched neuronal PRCs from Type II to Type I , inducinDifferences in network potentiation were especially pronounced for bability . Interesbability . On the bability  correspobability . Our simSince STDP requires correlated firing to potentiate the connection between two neurons, one might expect that asynchronous network activity should induce no net change in network potentiation, rather than the overall increased potentiation we observed. Further analysis of pre- and post-synaptic cell pairs uncovered an important statistical structure of the neuronal firing patterns in the cholinergically-modulated networks: post-synaptic neurons throughout the network were more likely to fire shortly after their pre-synaptic neurons rather than shortly before . Thus, pOn the other hand, the relatively lower network potentiation observed in networks with low cholinergic modulation was due to post-synaptic neurons firing right before their pre-synaptic partners much more frequently . This efIt should be noted that we tested this result for robustness against noise by adding Gaussian-distributed noise with a temporal correlation of 100 ms  to the external constant current driving individual neurons. We found that even for a noise amplitude as high as rameters . This noreached  .Since acetylcholine levels vary dramatically in cortex, we investigated how sensitively our results depended upon acetylcholine levels, which dramatically influence PRC shape. Cholinergic modulation was modeled by varying the slow potassium conductance We also tested our results for robustness to connectivity density by increasing the radius of connectivity in our network simulations . High-ACh networks showed greater overall potentiation than low-ACh networks for a wide range of connectivity densities (0.8% to 4.0% connectivity), though sparser connectivity led to greater differences in network potentiation .We tested the results for robustness to frequency modulation by varying the duration of the STDP window, Finally, several studies have shown that the equilibrium distribution of synaptic weights in a network subject to STDP strongly depends upon the mathematical form of the STDP rule. For example, some have suggested that the integral of the LTD portion of the STDP curve should be greater than the LTP portion of the curve in order to maintain network potentiation at reasonable levels The above results pertained to networks with homogeneous connectivity distributions in the sense that all synapses could achieve the same maximal strength, and long-range network connections did not preferentially target any particular neurons. Such homogeneity certainly does not exist in the brain In this heterogeneous network, we alternately switched between the high and low acetylcholine concentration , and found that such switching induced immediate and dramatic changes in network synchrony and potentiation . As in t .This enhanced depotentiating effect resulted from the dynamical interplay between the cluster and the rest of the network. As shown in increases in strength during epochs when ACh is low, in contrast to the collective weakening exhibited by connections in the rest of the network.We have proposed a novel physiologically-plausible mechanism, based on cholinergic modulation of neural membrane excitability, that can account for synaptic renormalization during NREM sleep. We have shown that the dramatic changes in membrane excitability induced by cholinergic modulation, and the resulting changes in network firing patterns, lead to upscaling and downscaling of mean synaptic efficacy. Thus, our results propose a dynamical mechanism for synaptic renormalization that provides a bottom-up framework linking changes in the neuromodulator environment during waking and NREM sleep to changes in neuronal excitability, network activity patterns, and overall renormalization of network connectivity. Simulations of networks with heterogeneous synaptic connection distributions also provided evidence for selective rescaling of particular network connections.Our simulations showed that high levels of acetylcholine in cortical networks led to asynchronous dynamics, which in turn led to relatively high network potentiation. On the other hand, low levels of acetylcholine resulted in more synchronous network activity and relatively lower overall potentiation. These results are consistent with the prediction of the synaptic renormalization hypothesis that wakefulness (during which ACh is present at high levels in cortex) is associated with global synaptic upscaling, while NREM sleep (during which ACh is present at much lower levels in cortex) is associated with global synaptic downscaling. These results were also robust to noise, changes in network frequency, different network topologies, and various STDP parameters, and they were strengthened by network heterogeneities. Additionally, The desynchronization of neuronal activity that resulted from high concentration of ACh in our model is expected from PRC theory, since higher ACh induces more Type I-like PRC On the other hand, ACh is known to be down-regulated during NREM sleep, when slow wave activity dominates EEG recordings. Such activity is associated with the slow oscillation of thalamocortical neuron membrane potential that results from thalamocortical bistability While our theory focuses on possible dynamical underpinnings of the renormalization hypothesis, there are many other factors which may contribute to synaptic renormalization. Incoming sensory signals may promote upscaling during wakefulness Our theory hinges on the result that synchronous network activity leads to synaptic downscaling, while asynchronous network activity generates synaptic upscaling. Our analysis of the structure of spike times in pre- and post-synaptic cell pairs indicates that downscaling was due to timing competition between arriving excitatory post-synaptic potentials (EPSPs) within the brief period of synchronous spiking activity. This competition within such a short time window resulted in about half the pre-post pairings falling in the negative portion of the STDP curve and therefore leading to lower network potentiation relative to asynchronous network activity. It has previously been shown that asynchronous neuronal activity leads to increased network potentiation while synchronous activity leads to decreased network potentiation in simulated networks incorporating STDP with propagation delays In summary, we have shown that cholinergic modulation can lead to changes in overall network potentiation, and that these changes may be understood in terms of the altered cellular and network dynamics induced by ACh. Further experimental investigation into the possible role of cholinergic modulation in the dynamical underpinnings of synaptic renormalization is clearly required.The cortical pyramidal model neuron we employed was motivated by a recent experimental study which showed that in slices of mouse visual cortex, the presence of acetylcholine (ACh) modulated the response properties of cortical neurons as measured by the phase response curve (PRC) bustness . This noActivation of the The slow, low-threshold To obtain the phase response curves displayed in We simulated networks with 800 excitatory neurons and 200 inhibitory neurons. The network connectivity pattern was constructed using the Watts-Strogatz architecture for “small world networks” Synaptic current was transmitted from neuron depotentiated final synaptic distributions. All simulations were numerically integrated in Matlab using a fourth-order Runge-Kutta method with a time step of 0.05 ms.Simulations were initialized with all synaptic strengths set to any value, not just zero. The MPC between a pair of neurons, We quantified phase-synchronization of neuronal firing in our simulations using the mean phase coherence (MPC) measure In our simulations exploring network heterogeneity, the network was composed of 1000 neurons , of which 50 comprised a cluster in which


The ubiquitous challenge from infectious disease has prompted the evolution of diverse host defenses, which can be divided into two broad classes: resistance (which limits pathogen growth and infection) and tolerance . Resistance and tolerance may provide equivalent short-term benefits, but have fundamentally different epidemiological consequences and thus exhibit different evolutionary behaviors. We consider the evolution of resistance and tolerance in a spatially structured population using a stochastic simulation model. We show that tolerance can invade a population of susceptible individuals  with higher cost than resistance, even though they each provide equivalent direct benefits to the host, because tolerant hosts impose higher disease burden upon vulnerable competitors. However, in spatially structured settings, tolerance can invade a population of resistant hosts only with lower cost than resistance due to spatial genetic structure and the higher local incidence of disease around invading tolerant individuals. The evolution of tolerance is therefore constrained by spatial genetic structure in a manner not previously revealed by nonspatially explicit models, suggesting mechanisms that could maintain variation or limit the occurrence of tolerance relative to resistance. Parasites are ubiquitous and have detrimental effects on their hosts , resultiResistance and tolerance often incur physiological costs that manifest as reduced host fitness in the absence of disease in comparison to susceptible  hosts. Such costs can arise because the defense strategy has harmful pleiotropic effects  or becauResistance and tolerance traits, even given equivalent direct single-generation effects on host fitness , can have different epidemiological consequences and therefore exhibit fundamentally different evolutionary behaviors. By limiting pathogen growth and reproduction, resistant hosts can reduce further transmission of the pathogen and thus disease prevalence in the population. As resistant hosts increase in frequency relative to susceptible hosts, the fitness advantage of resistant hosts declines along with the decreasing probability of pathogen encounter. Such negative frequency-dependent selection is expected to maintain polymorphism in resistance over a wide range of costs . In contThere is growing recognition that the spatial structure of populations can markedly impact the evolution of disease-related traits. Theoretical  and empiHere we consider the evolution of resistance and tolerance with respect to their associated costs using a spatially explicit, individual-based stochastic simulation model. We assume that hosts exist on a regular lattice and that ecological interactions, such as pathogen transmission and host reproduction, can occur either locally between neighboring sites or globally between all sites on the lattice. Using pairwise invasion trials, we show that tolerance has an advantage relative to resistance when invading into a population of susceptible hosts. However, in a spatially structured setting, tolerance has a disadvantage relative to resistance when invading into a resident population containing well-defended resistant hosts. Our results show that spatial genetic structure can restrict the evolution of tolerance and extend our understanding of the relative advantages and costs of these forms of defense and the ease with which tolerance can evolve in host–parasite systems with spatial structure.R), tolerant (T), or susceptible (S). Resistance is defined here as the ability of the host to reduce the growth of the pathogen, thus reducing both the loss of host fecundity due to infection and the pathogen transmission from the infected host, but not the probability of becoming infected (avoidance). Tolerance is defined as the host's ability to offset the negative effects of infection on host fecundity, but without limiting pathogen growth or reproduction and thus without effects on pathogen transmission.We consider a regular 100 × 100 toroidal lattice of sites, which may each be empty or occupied by a host individual. Hosts may be of three asexual haploid genotypes, resistant  without affecting mortality. We checked that an otherwise identical model in which infection instead caused higher host mortality yielded qualitatively similar outcomes.Hosts may be infected by the pathogen and cannot recover from infection. Each host individual has a genotype Rr,1 = Tr,1 > Sr,1). In uninfected hosts, resistance and tolerance each incur a cost manifested as reduced fecundity of uninfected individuals relative to susceptible individuals . For convenience, we refer to the cost of defense Gc = 1 −ρG, where ρG = Gr,0/Sr,0 expresses the fecundity of uninfected resistant or tolerant individuals relative to uninfected susceptible individuals.To determine the evolutionary consequences of the different epidemiological consequences of resistance and tolerance, we considered resistance and tolerance traits that confer equivalent direct fitness benefits to infected individuals . Tolerance, in contrast, does not limit pathogen growth; thus, the transmission rate from infected tolerant hosts is equal to that from infected susceptible hosts . No pathogen transmission occurs from uninfected individuals.In our model, the fundamental difference between resistance and tolerance is in the pathogen transmission rate from infected individuals. Because resistance limits pathogen growth, the pathogen has lower transmission rate from infected resistant hosts than from infected susceptible hosts ; modifications to the model by weighting global transmission by the transmission rate β of each individual and by using iterations with global reproductive dispersal of the host as a means to disrupt local spatial structure gave qualitatively similar results. Lastly, individuals are removed by death at a fixed mortality rate μ.The model is iterated through synchronous time steps that consist of host reproduction, transmission, and death. First, host individuals reproduce into an adjacent empty site with a probability of success given by their fecundity Rc/T from 0 to 0.50 in increments of 0.02. In trials of the invasion of tolerance into resistant populations, we varied the cost of resistance Rc between 0 and 0.50 in increments of 0.02, while the cost of tolerance was varied in increments of 0.008 about a given cost of resistance. These particular values were chosen to allow reasonable resolution of cost differences within computational constraints. Invasion conditions were examined under both local and global transmission for each combination of costs.We studied the pairwise invasion of either resistant or tolerant hosts into susceptible populations, and the invasion of tolerant hosts into resistant populations. The complementary case of invasion of resistant hosts into tolerant populations was also examined and yielded outcomes qualitatively identical to the invasion of tolerant hosts into resistant populations (data not shown). In each case, we varied the relative costs of the defense traits to determine the effect of cost on invasion ability. Because the defense traits confer equivalent direct protective benefit, any differences in the relative costs at which resistance and tolerance can invade reflect differences in the overall fitness advantage provided by the defenses, accounting for their divergent epidemiological feedbacks. In trials of the invasion of resistance or tolerance into susceptible populations, we varied the cost of defense In each invasion trial, the lattice was initially populated with 1000 host individuals distributed randomly among sites, each with 0.25 probability of being infected. The simulation was run for 1000 time steps to allow the resident population to equilibrate. Invading hosts  were then introduced into random empty sites; infection prevalence among the invading hosts was set equal to that of the resident host population. The simulation was propagated for either 200,000 time steps or until one type of host reached fixation. Invasion dynamics were characterized by observing infection prevalence and the relative frequencies of the host types. Results from three independent replicates were averaged for each parameter combination.The parameter values that were explored are presented in T*c = 0.533; resistance, R*c = 0.529), suggesting that tolerance has an advantage relative to resistance during invasion of susceptible populations. Thus, our model replicates the prior result in unstructured susceptible populations that tolerance provides an advantage beyond the direct effects of protection from pathogen-induced fitness loss, in comparison to resistance . However, in spatially structured populations that contain only resistant hosts, tolerance can invade only if it has a lower cost than resistance that provides equivalent direct benefits  higher cost than resistance . We found that complete resistance and tolerance show different evolutionary behavior than partial defenses in the spatial setting. Complete tolerance can invade a population of susceptible hosts with substantially (up to 7%) higher cost than complete resistance , with the difference in costs comparable to that observed in the case of partial defense. Thus, complete tolerance has an advantage relative to complete resistance when invading susceptible populations. However, completely tolerant hosts can invade a population of completely resistant hosts with costs up to those of the resident resistant hosts, indicating that complete tolerance, unlike partial tolerance, suffers no disadvantage when competing with resistance that confers equivalent direct protective benefits.Hosts with complete resistance and tolerance defenses suffer no fitness loss due to infection (i.e., Resistance and tolerance each mitigate the harm caused by infectious diseases, but do so in ways that differ fundamentally in how they affect pathogen persistence and prevalence. In 2004, Restif and Koella suggested that tolerance favors the spread of pathogen and thus creates a harsh pathogenic environment for susceptible competitors. This effect of tolerance on competition among hosts represents an additional benefit that can allow tolerance to evolve with higher cost than resistance. Our results suggest that, on the one hand, this additional benefit of tolerance relative to resistance occurs when the defense arises in a population of susceptible host individuals that suffer relatively greater harm, in terms of fitness, due to disease. On the other hand, when tolerance is introduced into populations composed of equally well-defended resistant hosts, the increased pathogen burden does not harm the resistant hosts any more than it harms the tolerant hosts themselves; thus, no additional competitive advantage is afforded to tolerance. Our results therefore suggest that the presence of resistance can potentially limit the evolution of tolerance.Many natural systems exhibit spatial genetic structure over short distances due to limited dispersal of gametes or offspring . In our Unlike hosts that have partial defenses, hosts with complete defenses, by definition, suffer no fitness loss due to infection. Completely tolerant hosts therefore experience no self-imposed fitness burden due to locally elevated infection prevalence, yet, still benefit from imposing disease-induced harm upon susceptible competitors. Consequently, complete tolerance can invade susceptible populations with costs greater than those of complete resistance and can invade resistant populations with costs up to those of resistance.While epidemiological feedbacks have been recognized as an important driver of the evolution of defenses , previouGiven that disease resistance is widespread in natural populations, the introduction of tolerance by either mutation or migration probably often occurs in populations that contain resistant hosts. Our findings suggest that barriers to the evolution of tolerance within spatial settings may be greater in the presence of resistance. This may contribute some understanding to the puzzling observations of variation in tolerance within and among populations , 2009, dThe main driver of the relative fitness of resistant and tolerant genotypes is the relative amount of pathogen transmission from infected hosts. Mixed strategies of defense are therefore likely to exhibit distinct evolutionary dynamics . For giv


Non-enzymatic protein glycation is a source of metabolic stress that contributes to cytotoxicity and tissue damage. Hyperglycemia has been linked to elevation of advanced glycation endproducts, which mediate much of the vascular pathology leading to diabetic complications. Enhanced glycation of immunoglobulins and their accelerated vascular clearance is proposed as a natural mechanism to intercept alternative advanced glycation endproducts, thereby mitigating microvascular disease. We reported that antibodies against the glycoprotein KLH have elevated reactivity for glycopeptides from diabetic serum. These reactions are mediated by covalent binding between antibody light chains and carbonyl groups of glycated peptides. Diabetic animals that were immunized to induce reactive antibodies had attenuated diabetic nephropathy, which correlated with reduced levels of circulating and kidney-bound glycation products. Molecular analysis of antibody glycation revealed the preferential modification of light chains bearing germline-encoded lambda V regions. We previously noted that antibody fragments carrying V regions in the germline configuration are selected from a human Fv library by covalent binding to a reactive organophosphorus ester. These Fv fragments were specifically modified at light chain V region residues, which map to the combining site at the interface between light and heavy chains. These findings suggest that covalent binding is an innate property of antibodies, which may be encoded in the genome for specific physiological purposes. This hypothesis is discussed in context with current knowledge of the natural antibodies that recognize altered self molecules and the catalytic autoantibodies found in autoimmune disease. The generation of an enormous diversity of antibodies in response to the multitude of possible antigens is a signature of instructive or adaptive immunity. The structural basis for adaptive immunity is expressed in the variability of the antigen binding sites displayed on antibodies and B cell receptors. Thus, antibodies are conventionally associated with the genetic recombination and accumulated mutations in their variable (V) regions that incrementally improve the complementarity between the antibody combining site and groups on the antigen. In contrast to affinity that matures gradually over time through multiple weak interactions, binding through strong forces such as a covalent bond could enable a more rapid and efficient way to capture certain antigens. Is there any case where antibodies use this form of binding and what purpose could such a binding mechanism serve?Antibodies that bind ligands covalently have been sought in approaches to generate enzyme-like catalytic antibodies . CovalenIn an alternative framework one could postulate that covalent binding antibodies might also occur naturally if this activity were advantageous to the host. We proposed that binding through a single strong interaction to an antibody would be an appropriate mechanism for the sequestration and clearance of chemically damaged proteins and cells. Such a function is increasingly recognized in studies of naturally occurring antibodies that have inherent affinity for altered structures on self . For exaAnother kind of cytotoxic metabolic waste is generated through glycation or glycoxidation as sugars and carbohydrates are constantly bathing proteins and cells and modifying them through nonspecific reactions of their exposed carbonyl groups. Glycation is a slow and continuous process that occurs in normal aging. However, it is significantly elevated in diabetes due to recurring hyperglycemia or poor glycemic control. The role of this pathway in leading to vascular complication of diabetes is now firmly established -10. A beA favorable chemical reaction of antibodies with glycation products is suggested by the enhanced glycation of normal immunoglobulins -24. GlycIn order to test this hypothesis we sought to demonstrate that covalent binding antibodies could be elevated in an immune response and that these could attenuate a glycation stress by reducing the carbonyl load. Reactive immunization suggested a feasible approach. Previously, we showed that antibodies induced to a pyruvate-containing hapten-KLH conjugate could bind to antigen by recognizing only the carbonyl group in the hapten . HoweverThe streptozotocin-induced diabetic rat, serving as a suitable model for diabetic nephropathy, was used to demonstrate the potential of the reactive immune response to mitigate AGE formation in vivo. Compared to diabetic animals immunized with adjuvant alone, KLH-immunized diabetic rats had significantly diminished AGE levels, which correlated with attenuated nephropathy . ImproveThese studies provide evidence that a natural covalent reactivity of antibodies is augmented by adaptive immunity. However, the predominance of unmutated V region sequences in the reaction products suggested that reactivity is inherent to the germline encoded VL . How couIn an alternative approach, we used synthetic reactive substrates as antigens to probe for antibodies capable of covalent binding. This idea followed the original concept that antibodies selected for nucleophilic reactivity could also express enzymatic activity through covalent catalysis. Irreversible covalent binding to a small organophosphorus (OP) ester was used to chemically select single-chain VH-VL fragments (scFv) from a phage display library . All of Chemical selections in vitro might represent the first step in adaptive immunity for acquisition of reactivity. However, the contribution of irreversible covalent binding to clonal selection in vivo remains speculative. Using a reactive OP esterprotein conjugate as immunogen, we examined the potential for the immune response to elicit reactive antibodies that are modified by the antigen. Antisera against the OP conjugate included reactive antibodies, which could be detected by their covalent modification with biotin-tagged OP reagent . RemarkaIgM nAbs are predominantly produced by a population of long-lived, self-replenishing B-cells, including the B-1 and B-1a subsets. It is believed that this B cell repertoire is conserved in evolution for its contribution to host defense . The gerOxidized phospholipids, which accumulate in atherosclerotic lesions, are targets of both innate and adaptive immunity . The rolPolyreactivity of nAbs is defined as the promiscuous avidity of multivalent IgM molecules to disparate molecules, including intracellular proteins and nucleic acids . This biReports showing that DNA autoantibodies found in lupus or other autoimmune diseases have phosphodiesterase activity suggests a further case where molecular recognition in the Ig V domains is manifested as chemical reactivity . These aCovalent-binding antibodies obtained by immunization or reactive selection and catalytic antibodies found in autoimmune pathologies appear to have features in common with nAbs. These features include VH and VL domains in germline or near-germline configurations and the recognition of molecules presenting uncommon chemical functionality, such as reactive carbonyls and phosphorus esters. Such features may be fundamentally related in that the chemical or enzymatic function is presumably highly evolved and therefore conserved in the genome. Whereas nAbs are IgMs that rely on weak multivalent binding to molecular patterns on antigens for high avidity, chemically reactive or catalytic antibodies are also expressed as IgG that can bind antigens with high affinity. However, the latter may also react with diverse substrates through weak, noncovalent pre-reaction complexes. According to the transition state theory of catalysis the strongest binding is expressed between an enzyme and the transition state. In the covalent complex, binding is dominated by the chemical bond between a residue of the active site and a group on the ligand. The focus of the weak binding by nAbs on small, minimally altered epitopes is consistent with the recognition of substrate functionality for reactivity. Thus, chemical reactivity could provide a mechanism to translate weak non-covalent binding to strong binding or to catalytic activity. The physiological purpose of chemically reactive and catalytic antibodies must also be addressed, even as the role of nAbs in protective immunity and homeostasis is only beginning to emerge. Participation of nAbs as response elements in oxidative stress and apoptotic cell clearance suggests a housekeeping function that predates the evolution of adaptive immunity. Similar considerations may apply to the rationale for covalent-binding antibodies acting as buffers against glycation stress. The preservation of covalent reactivity or catalytic functions in adaptive responses and in autoimmunity could be attributed to biological design or to defects in immune regulation. Inducibility through adaptive immunity might offer an appropriate mechanism to invoke beneficial responses to oxidative or glycoxidative stress. On the other hand, the role or contribution of natural catalytic autoantibodies in autoimmune pathology remains obscure, and their existence could simply reflect a failure in the V gene repertoire shift on induction of IgG autoantibodies , 65. DesThis work was supported by the American Diabetes Association grant 1-05-RA-136, NIH grant CA90564, and the UC Davis, Medical School - Children's Miracle Network.


The incidence of obesity and overweight has reached epidemic proportions in the developed world as well as in those countries transitioning to first world economies, and this represents a major global health problem. Concern is rising over the rapid increases in childhood obesity and metabolic disease that will translate into later adult obesity. Although an obesogenic nutritional environment and increasingly sedentary lifestyle contribute to our risk of developing obesity, a growing body of evidence links early life nutritional adversity to the development of long-term metabolic disorders. In particular, the increasing prevalence of maternal obesity and excess maternal weight gain has been associated with a heightened risk of obesity development in offspring in addition to an increased risk of pregnancy-related complications. The mechanisms that link maternal obesity to obesity in offspring and the level of gene-environment interactions are not well understood, but the early life environment may represent a critical window for which intervention strategies could be developed to curb the current obesity epidemic. This paper will discuss the various animal models of maternal overnutrition and their importance in our understanding of the mechanisms underlying altered obesity risk in offspring. The current epidemic of obesity and related metabolic disorders has been seen as a symptom of affluence with the primary cause relating to the development of an obesogenic environment and ease of access to highly calorific foods and reduced energy expenditure in work and leisure activities . The metin utero, they may be maladaptive in later life, contributing to adverse health outcomes when offspring are exposed to catch-up growth, diet-induced obesity, and other factors [The mechanisms underpinning the developmental programming framework and the role of genetic versus environmental factors remain speculative. One general thesis is that in response to an adverse intrauterine environment the fetus adapts its physiological development to maximize its immediate chances for survival. These adaptations may include resetting metabolic homeostasis set points, endocrine systems, and downregulating of growth, commonly manifest in an altered birth phenotype. More recently the “predictive adaptive response” (PARs) hypothesis proposes that the degree of mismatch between the pre- and postnatal environments is a major determinant of subsequent disease risk , 8. Thus factors . mechanisms underlying phenotype development [Animal models have been extensively used to study the basic physiological principles underlying the DOHaD hypothesis and are essential to the search for the mechanistic links between prenatal and postnatal influences and risk for developing the metabolic syndrome in later life. Epidemiological data suggest that developmental programming occurs within the normal range of birth size , 11, butelopment . This paper se, induces hyperleptinemia and insulin resistance in offspring, as well as an increased body weight that persists into adulthood [The rodent is the most commonly used model species for investigation of developmental programming via a maternal obesogenic nutritional environment. A maternal cafeteria or high-fat (HF) diet has been shown to induce obesity, insulin and leptin resistance –17, hypedulthood . Even midulthood . Our owndulthood   and suppressor proopiomelanocortin (POMC) in offspring at weaning . Howevere levels .γ [Work by Simar et al. in the adult rat revealed an interaction between maternal obesity and postnatal overnutrition on skeletal muscle metabolism, a postweaning HF diet exerted an additive effect to that of maternal obesity on body weight and skeletal muscle markers of glucose and lipid metabolism but not on plasma glucose and insulin levels, suggesting that maternal obesity and postnatal overnutrition impair skeletal muscle function via different mechanisms . Reducedγ .Although several studies have examined changes in locomotor activity in the setting of maternal undernutrition –63, dataAltered placental function in the setting of maternal obesity has also been the focus of several investigations. Our group and others have reported altered placental structure and function as a result of a maternal HF diet across a range of experimental models. In the pregnant rat, a maternal HF diet has been shown to reduce growth of the fetus and the placental junctional zone, but not placental labyrinth zone growth . In the Until relatively recently, developmental programming was seen as an irreversible change in developmental trajectory. Outside of the early taurine reversal work in the setting of the maternal low protein model , there iThere is evidence for a role of diet reversal in ameliorating the effects of maternal obesity on offspring outcome. In the nonhuman primate, diet reversal from an HF to control diet during pregnancy led to normalisation of the melanocortin levels, improvements in fetal hepatic triglycerides, and partial normalization of the expression of gluconeogenic enzymes. These results suggested that simply changing to a normal low-fat diet, specifically during pregnancy, can lower, but not eliminate, the risk for fetal hepatic steatosis . SimilarEpigenetic processes lead to heritable changes in gene function by altering DNA chemistry independent of sequence and may be responsible for tissue-specific gene expression during differentiation. Epigenetic modifications may be one mechanism by which exposure to an altered intrauterine milieu or metabolic perturbation may influence the phenotype of the organism much later in life . HoweverExperimental data in rodents and recent observations in humans suggest that epigenetic changes in regulatory and growth-related genes play a significant role in mediating the pathophysiological phenotypes derived from developmental programming , 72. Hisα [in vitro is prolonged [Adversity during pregnancy or early neonatal life in experimental programming models results in changes in promoter methylation, therefore, directly or indirectly, affect gene expression in pathways associated with a range of physiologic processes . For exaα , 75, infα  into posα , 78. Manrolonged .It has been shown that the promoter in the leptin gene is subject to epigenetic programming, and leptin gene expression can be modulated by DNA methylation –82. ReceDevelopmental epigenetics is believed to establish “adaptive phenotypes” to meet the demands of the later-life environment , 87. Impβ-cell “dysfunction” in rat F1 female offspring. Paternal HF diet altered the expression of 642 pancreatic islet genes in adult female offspring including functional gene clusters related to cation and ATP binding, cytoskeleton, and intracellular transport. Broader pathway analysis demonstrated involvement of calcium-, MAPK-, and Wnt-signalling pathways, apoptosis, and cell cycle regulation. Hypomethylation of the Il13ra2 gene, which showed the highest fold difference in expression, was demonstrated. This work provided the first evidence in mammals of nongenetic, intergenerational transmission of metabolic sequelae of an HFD from father to offspring [There is now evidence supporting a role for paternal transmission of disease risk across generations. Ng et al. recently reported that a paternal high-fat diet can program ffspring . Yazbek ffspring . Of noteffspring  which suAnimal studies across a range of species and varied obesogenic diets have provided clear evidence linking maternal obesity and increased risk for obesity in offspring and are invaluable tools in the investigation of mechanisms underpinning this linkage. Given the current obesity epidemic and the increasing number of obese women entering pregnancy, there is an urgent need for interventional strategies that target developmentally programmed obesity. The evidence available on short- and long-term health impact for mother and child currently favours actions directed at controlling prepregnancy weight and preventing obesity in females of reproductive ages  although


Clinical observations suggest abnormal gaze perception to be an important indicator of social anxiety disorder (SAD). Experimental research has yet paid relatively little attention to the study of gaze perception in SAD. In this article we first discuss gaze perception in healthy human beings before reviewing self-referential and threat-related biases of gaze perception in clinical and non-clinical socially anxious samples. Relative to controls, socially anxious individuals exhibit an enhanced self-directed perception of gaze directions and demonstrate a pronounced fear of direct eye contact, though findings are less consistent regarding the avoidance of mutual gaze in SAD. Prospects for future research and clinical implications are discussed. Social anxiety disorder (SAD) is one of the most common mental disorders with a lifetime prevalence of up to 12% in Western countries  whether mutual gaze is more readily perceived; and (b) whether mutual gaze is avoided and perceived as threatening.Most mammals generally interpret direct gaze as threatening or as a sign of dominance. Humans in contrast often associate mutual gaze with positive interest, such as love and attraction. A preference for direct gaze seems to be present at a very early age: Farroni et al.  found thDifferent sources of information are taken into account when processing gaze direction. The most obvious cue lies in the eye itself. Kobayashi and Kohshima , 2001 coAnother factor influencing gaze perception is the head direction of the looker. Langton  see als found thStudies that focus on the ability to distinguish between direct and averted eye gaze are relatively numerous. All these studies generally report that human observers are highly accurate at determining mutual eye gaze. In their classic study, Gibson and Pick  asked obFacial emotional expression is another cue taken into account when judging gaze directions. Lobmaier et al.  presenteEwbank et al.  employedThe studies reviewed above reveal that gaze perception plays an important role in social interactions and is modulated by several factors, such as head direction, interpersonal distance, or emotional facial expressions  higher amplitudes of the P100 in response to averted gaze  and is followed by attentional avoidance of such stimuli (e.g., Wieser et al., Measuring scan paths by means of eye-tracking is a highly ecologically valid method to assess overt gazing behavior. Hence, eye-tracking methods seem highly suitable to study avoidance of mutual gaze in individuals with social anxiety. In addition, such methods also allow studying approach-avoidance behavior in response to more ecologically valid stimuli, such as films or crowds of individuals (Lange et al., Further research is needed to assess the diagnostic value of abnormalities in gaze perception as possible behavioral indicators of SAD. To date, statements regarding the diagnostic potential of such measures are substantially limited since none of the studies included a clinical comparison group, comprising for instance individuals with symptoms of autism, or schizophrenia who also exhibit abnormal gaze perception (Kliemann et al., In sum, recent findings highlighted abnormal gaze perceptions in social anxiety. In particular, socially anxious individuals were characterized by a greater self-referential perception of gaze direction along with a pronounced fear of direct eye contact.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.


Vireo atricapilla) suffered a substantial contraction of breeding habitat and population size during much of the 20th century. In a previous study, we reported significant differentiation between remnant populations, but failed to recover a strong genetic signal of bottlenecks. In this study, we used a combination of historical and contemporary sampling from Oklahoma and Texas to (i) determine whether population structure and genetic diversity have changed over time and (ii) evaluate alternate demographic hypotheses using approximate Bayesian computation (ABC). We found lower genetic diversity and increased differentiation in contemporary samples compared to historical samples, indicating nontrivial impacts of fragmentation. ABC analysis suggests a bottleneck having occurred in the early part of the 20th century, resulting in a magnitude decline in effective population size. Genetic monitoring with temporally spaced samples, such as used in this study, can be highly informative for assessing the genetic impacts of anthropogenic fragmentation on threatened or endangered species, as well as revealing the dynamics of small populations over time.Anthropogenic alterations in the natural environment can be a potent evolutionary force. For species that have specific habitat requirements, habitat loss can result in substantial genetic effects, potentially impeding future adaptability and evolution. The endangered black-capped vireo ( Among the challenges to understanding evolution at the population level is unveiling demographic history using genetic data. Although ‘snapshots’ of genetic patterns based on single samples can be extremely informative, the ability to divulge historical population processes that shaped current patterns of genetic variation is limited . This caThis need to understand the potential evolutionary outcomes of past demography is nowhere as crucial, perhaps, as in cases where the genetic impacts of anthropogenic actions need to be assessed – as is often the need in conservation studies. For species existing in small populations and those of conservation concern, it may be necessary to unravel demographic history to evaluate whether genetic sustainability is of concern , and to Vireo atricapilla) – an endangered bird species.One avenue for circumventing the above limitations is to use archived specimens and molecular techniques sufficiently sensitive for working with limited or degraded DNA . Such saMolothrus ater), associated with increasing habitat edges, are considered as important influences. Severe parasitism by cowbirds is considered to be a relatively recent problem – having increased together with fragmentation – although it is reasonable to expect that some parasitism has always existed.The black-capped vireo is a neotropical songbird that breeds in the early-successional oak-juniper savanna ranging from south-central Texas through southern Oklahoma (USA). This species is reported to have suffered substantial losses in both its breeding and wintering habitat because of agriculture and urbanization over the past 50 years . At the Recently, we reported that many populations of black-capped vireos showed more structure than might be expected, based on the vagility of these birds . There wNev) to understand the impact of drift on remnant populations.Little is known about the demographic history of this species, and especially about past genetic diversity and population sizes. In this study, we (i) compare diversity indices and population structure from historical and contemporary populations, (ii) use approximate Bayesian computation models to explore the demographic history – especially with regard to the amplitude and timing of putative population size changes and (iii) estimate long-term effective population size (We focused on the availability of multiple samples collected from 1899 to 1915 at locations from areas corresponding with the contemporary samples collected during 2005–2008. We chose this period because (i) few samples existed in collections either before or after this time from multiple locations and (ii) the sampling straddles the most likely period of habitat and population decline. Three locations  represenContemporary samples were collected in 2004–2006, with additional samples collected in 2007 and 2008. Methodology of sample collection is similar to the one described by 3 patch of skin removed from the toe-pads of museum specimens using sterile surgical razors and then placed in a sterile microcentrifuge tube  using GENETIX (AR), which controls for biases in allelic diversity arising from unequal sample sizes  to test the hypothesis that the sample medians do not between differ time points. For historical comparisons, where we have three time points, we used a Kruskal–Wallis test , blocked by locus, to compare HEXP and AR among sampling periods.To estimate changes in genetic diversity between a) historical and contemporary samples or b) between pairs of samples within the contemporary period, we used a Wilcoxon signed-rank test . This nonparametric analysis uses estimates of FST (Θ of Weir and Cockerham) among the 1905–1910 historical samples from Kerr, Bexar and Oklahoma populations in GENETIX. FST estimates were obtained using 3000 permutation resampling to control for possible variation because of differences in sample sizes. We performed identical analyses on the corresponding three contemporary populations for comparison. We used a Wilcoxon signed-rank test (paired by locus) to see whether the degree of differentiation among historical samples was different from that of contemporary samples, for the three population pairs. Pairwise differentiation values for each locus were the replicates in the statistical comparison. Using FST to infer differentiation in cases where intra-population diversity is high can be problematic  may have taken place and to estimate the historical and present population sizes. Coalescent inference methods can utilize genetic summary statistics to extract evolutionary histories , and theblematic .Ne and timing of the putative decline for the whole species and (ii) incorporating divergence events among sample sites in the coalescent genealogies would exclude gene flow between lineages, which we know to be incorrect.The ABC approach can be briefly summarized as follows: First, coalescent population gene genealogies are simulated from a uniform prior distribution for each parameter of interest. Priors are defined based on what is known from the species biology, or based on estimates for similar species. Then, genealogy and parameter space is explored through Markov chain Monte Carlo (MCMC) algorithms with a range of uniform priors, producing a large reference table of summary statistics from which genetic data are simulated. Second, a rejection algorithm using We coded 11 potential scenarios to explore demographic history and to detect bottlenecks from our two temporal samples, and a total of 11 million data sets were simulated. A summary of scenarios evaluated and priors used are presented in Ne in our sampled populations. For microsatellite data, the temporal method is reported to be more precise when samples are taken several generations apart than point estimates , where it is assumed that migrants originate from an infinitely large population. This was important because, despite our finding of population differentiation, FST values indicate that the study populations experience gene flow  population by combining samples from the other two remaining populations. While the assumption (infinitely large source) is typical for many idealized populations, in reality few species satisfy this assumption. Although historical populations may not have been infinite, they may have represented a panmictic population, without significant population structure. Hence, we avoided using contemporary populations as the source population as gene flow from differentiated populations may produce biased estimates , we assumed three potentially different generation lengths  to evaluate the effects that T may have on estimates of Ne. There is some variation among sexes (Concurrent estimation of stimates . Temporang sexes  regardinNe assuming closed populations using three approaches: (i) the moment estimator of Ne (MtNe) based on equations 15 and 18 from Ne) implemented in the program CoNe  and heterozygosity (HEXP) among the 1905, 1910 and 1915 samples from Kerr  being generally higher than FST and GST for both the historical and contemporary periods.Based on rom Kerr . In contulations . We alsoulations , with pontiation , despitet90 to N8 [200:1000] at t0  generations ago  for the Bexar and Oklahoma samples based on two sampling periods (1910–2005) were approximately 38  and 31 , respectively (The MLectively . For KerNe) produced the smallest estimates, coalescent estimator (CoNe) produced the largest estimates, and the likelihood estimator  produced estimates intermediate to the other two methods  for the historical population, and 872  for the modern population samples.Results from ONeSAMP gave HEXP followed a similar pattern as AR, although the proportional change was smaller in the former. Our observation of lower heterozygosity in contemporary relative to the historical populations indicates that inbreeding or genetic drift had occurred , drastic demographic changes appear unlikely during 1900–1915. Allelic richness was higher in the historical samples compared to the contemporary samples, indicating a population contraction. Nine alleles were not detected in the contemporary samples compared to historical samples. No reciprocal condition of unique alleles in contemporary populations was detected. Allelic richness is considered more sensitive to population bottlenecks than is heterozygosity . Estimatoccurred , b, proboccurred  as it taoccurred . AlthougThis historical divergence between Texas and Oklahoma populations may represent a long established discontinuity in vireo habitat that pre-dated urbanization of the mid-20th century. Historical vireo distributions in Texas and Oklahoma are separated by approximately 400 km  of unsuiThe Origin of Species (1859) is particularly prescient and relevant to our study: ‘Climate plays an important part in determining the average numbers of a species, and periodical seasons of extreme cold or drought, I believe to be the most effective of all checks. I estimated that the winter of 1854–1855 destroyed four-fifths of the birds in my own grounds; and this is a tremendous destruction, when we remember that 10% is an extraordinarily severe mortality from epidemics with man. The action of climate seems at first sight to be quite independent of the struggle for existence; but in so far as climate chiefly acts in reducing food, it brings on the most severe struggle between the individuals, whether of the same or of distinct species, which subsist on the same kind of food’.Coalescent approaches have become useful tools for uncovering important demographic events in the history of populations . In our While pulses of freakish climate change can undoubtedly spike mortality, the genetic and evolutionary consequences of drawn-out climate change will need to be investigated – especially when such changes are also occurring concurrently with anthropogenic habitat changes. It has been argued that the severity of the 1930s Dust Bowl was exacerbated by human land use changes over the preceding 50 years , which iNe approach assuming migration produced smaller estimates and wider confidence intervals compared to other estimators that assumed closed populations. It is important to note that the MLNe joint estimator assumes that rates of migrants are from an infinitely large population, gene flow remains constant over the sampling interval, or there is no significant subdivision among source populations. We now know from analysis of structure that these assumptions are not satisfied. However, most natural populations do not persist with constant migration rates over time (Ne. This offers an informative contrast with the approaches assuming closed populations. Bias has been a major concern with estimation of Ne based on temporal variance. However, these biases are expected to be less problematic with larger sample size, multiple sampling points, increasing number of variable loci and increasing the interval between samples. Ne from Kerr, for which population we have the best sample representation. ONeSAMP results also confirmed the trend that we observed with the ABC analysis, although actual estimates were higher for both the historical and contemporary samples.The MLver time . NonetheOverall, our results from the various approaches discussed here, such as decreases in allelic diversity, heterozygosity within populations, increased differentiation among populations and evidence for a demographic bottleneck based on coalescent simulations, suggest that a population decline of considerable effect has occurred between the historical and contemporary period. Furthermore, these demographic changes coincide with documented environmental and habitat changes that have occurred over the last century. Based on these results, a strong case can be made for the need to improve genetic connectivity between remnant populations, if adverse genetic consequences or loss of adaptive potential are to be prevented. Further, the steep trajectory of genetic decline indicates that this species may have a diminished buffering capacity against continued onslaughts on their habitat.Our work presented here provides a broadly applicable approach to address questions about population genetic change in cases where determining anthropogenic impacts may be valuable to future management. While use of archived genetic material is by no means novel, advances in molecular and computational methods make the use of limited archived tissue viable and especially valuable from a conservation perspective. We recently applied this approach successfully in revealing potentially critical details about the recent demography of another endangered species, the golden-cheeked warbler . This woHere, we were able to trace the genetic consequences of demographic change based on changes in allele frequencies over time, and by modeling alternative evolutionary scenarios. The ability to infer demographic history allows us to view genetic change in the context of habitat loss and fragmentation, and prognosticate on how this genetic change may impact the future persistence of populations.Black-capped vireo populations are rebounding from their low census numbers following intensive cowbird control and active management of habitat, including prescribed burns, to suit their habitat requirements . As a teAlthough the northern range of the species has contracted, it is believed that the breeding range extends southward farther than previously thought . Our stuOther similar studies  have shoQuerying the past with ancient or archived DNA provides information that helps resolve details of the otherwise hazy evolutionary past of populations or species. Inferences from such analyses allow us to evaluate which events of the past may be most consequential for the evolutionary future of populations.


My laboratory studies how chromosomes are organized in three dimensions. The three-dimensional organization of the genome is critical for regulating gene expression by bringing genes in close spatial proximity to distal regulatory elements such as enhancers. We have developed powerful molecular approaches, based on our Chromosome Conformation Capture technology, to determine the folding of genomes at unprecedented resolution (Kb) and scale (genome-wide).We have applied these methods to determine the spatial folding of 1% of the human genome (the ENCODE pilot regions) across a panel of cell lines. We discovered that chromosomes fold into extensive long-range interaction networks in which genes are interacting with distal gene regulatory elements. These results start to place genes and regulatory elements, that are often separated by large genomic distances, in three-dimensional context to reveal their functional relationships.Our analysis of chromosome folding also revealed that chromosomes are compartmentalized in a series of “Topological Association Domains” (TADs) that are hundreds of Kb in size. Loci located within a TAD mingle freely, but interact far less frequently with loci located outside their TAD. TADs appear involved in gene expression, as we found that genes located within the same TAD tend to be co-expressed, but the mechanism(s) by which these domains affect gene regulation is still unknown. TADs represent novel universal and genetically encoded building blocks of chromosomes.


An earlier review summarized evidence relating use of snus (Swedish-type moist snuff) to health and to initiation and cessation of smoking. This update considers the effect recent publications on snus use and health have on the overall evidence. The additional evidence extends the list of neoplastic conditions unassociated with snus use  to include colorectal cancer and acoustic neuroma, and further undermines the weakly-based argument that snus use increases the risk of pancreatic cancer, although there is a report of poorer cancer survival in users. It remains undemonstrated that “snuff-dipper’s lesion” increases risk of oral cancer, and recent publications add to the evidence that snus use has no effect on periodontitis or dental caries. Although onset of acute myocardial infarction is not adversely associated with snus use, there is some evidence of an association with reduced survival. Whether this is a direct effect of snus use or a result of confounding by socioeconomic status or other factors requires further investigation, as does a report of an increased risk of heart failure in snus users. Even if some adverse health effects of snus use do exist, it remains clear that they are far less than those of smoking. Swedish-type moist snuff (“snus”) consists of finely ground air- or sun-cured tobacco, salt (sodium chloride), water, humidifying agents, chemical buffering agents (sodium carbonate), and food-grade flavourings. It is manufactured using a heat treatment process and, because of the manufacturing methods and selection of snus tobaccos, has stable levels of certain unwanted constituents, such as tobacco-specific nitrosamines, which are low compared to smokeless products prevalent on the US market, such as US-style moist snuff ,2. A pinIn recent years there has been considerable interest in snus as a possible safer alternative to smoking. However, attention has been drawn to possible health effects of snus use. A review in 2008 for the EC  referredSince my comprehensive review  appearedMethods are as previously described , with seNine new relevant publications were identified -27.An earlier review of smokeless tobacco and risk of pancreatic cancer  considerWhile there are no new studies on pancreatic cancer and snus, attention should be drawn to a recent paper  reportinOne must conclude that the evidence for an association of snus with pancreatic cancer, already weakly based before the pooled analysis , has becThe direct epidemiological evidence previously considered  found noA recent paper  describeA recent study  is the fAt the time of my earlier review , there wA recent paper  concernsTable four of my earlier review  summarizA recent paper  describeWhile these results seem consistent with a lack of effect of snus use on AMI, it should be noted that the pooled analysis , though Results relating to the short-term fatality rate were not presented by study, but it seems clear that they largely depended on the findings from the Construction Workers Study which contributed 88% of the total person-years of follow-up. The results seem consistent with an earlier report from this study  which reAnother recent relevant publication  investigThe earlier review  briefly These conclusions are supported by two recent publications ,27 basedBased on my previous review , I conclWhile this update does not consider evidence relating snus use to initiation or cessation of smoking, it does provide further information on possible health effects associated with snus use. As regards neoplastic conditions, recent evidence extends the list of those unassociated with snus use to include colorectal cancer  and acouNew publications on periodontitis  or dentaAlthough the evidence remains clear that effects of snus use on risk of CID, if they exist, are much less than those of smoking, recent publications have provided evidence of increased risk in snus users of heart failure  and of cAdverse health effects of snus use have not been clearly identified. The recent literature strengthens the evidence that any cancer risk (including that of pancreatic cancer) is at most minimal, and certainly much less than that associated with smoking. However, it does suggest that snus use might adversely affect survival from cancer. Although there is little evidence that snus use is associated with onset of AMI or stroke, reports of an increased risk of fatal AMI require further investigation.AMI: Acute myocardial infarction; BMI: Body mass index; CI: Confidence interval; CID: Circulatory disease; IHD: Ischaemic heart disease; MONICA: Monitoring of cardiovascular disease; OR: Odds ratio; RR: Relative risk.The author is a long-term consultant to the tobacco industry. However, this is an independent scientific assessment, the views expressed being those of the author alone.PNL was responsible for planning the study and writing the paper.


Plants exposed to abiotic stress respond to unfavorable conditions on multiple levels. One challenge under drought stress is to reduce shoot growth while maintaining root growth, a process requiring differential cell wall synthesis and remodeling. Key players in this process are the formation of reactive oxygen species (ROS) and peroxidases, which initially cross-link phenolic compounds and glycoproteins of the cell walls causing stiffening. The function of ROS shifts after having converted all the peroxidase substrates in the cell wall. If ROS-levels remain high during prolonged stress, OH°-radicals are formed which lead to polymer cleavage. In concert with xyloglucan modifying enzymes and expansins, the resulting cell wall loosening allows further growth of stressed organs. Abiotic stress in plants refers to growth conditions which are unfavorable for growth. Traditionally, these conditions include temperature stress (either cold or heat) as well as drought, osmotic stress and salinity. The latter stress factors are tightly associated with scarce water resources, a major challenge for future farming given the predicted increase of human population in the next decades  that is strongly induced by several abiotic stresses. They overexpressed the gene in tobacco plants and found a highly elevated peroxidase enzyme activity in the cell wall leading to transgenic plants which are more tolerant against salt and drought stress.Cell walls contain a number of structural proteins, as well as hemicellulosic polymers with attached phenolic residues like ferulated arabinoxylans, which can be cross-linked by the action of cell wall peroxidases  in plants, as a co-substrate. ROS have been observed to be a typical plant response to osmotic stress  constitutively in Arabidopsis, which was previously identified as an abiotic stress  induced gene in hot pepper. The strong reduction in root length observed in control seedlings of Arabidopsis on salt-containing media is less pronounced if the CaXTH gene is expressed. However, aberrant leaf morphology (curled leaves) was found in several independent lines and transverse sections of leaves showed an irregular cell pattern compared to wild type plants. The same CaXTH gene was later expressed in tomato plants without phenotypical side effects  mediated cell wall remodeling is based on a widely proposed cell wall model, in which xyloglucans act as tethering polymers between load bearing cellulose fibrils Hayashi, . The mosxxt1/xxt2 in Arabidopsis, which lacks classical xyloglucan due to a disruption of both xyloglucan xylosetransferases (XXT) genes  in tobacco. Roots of overexpressor lines developed far better under high salt conditions than wild type tobacco seedlings. It was suggested that seedlings with increased expansin expression have a higher water retention ability, though the possible mechanism as well as the potential change in the cell walls remain unclear. The overexpression of expansin A4 from rose in Arabidopsis leads to higher germination rates under salt stress, longer roots and an increase of lateral roots   growth, though with a reduced rate. Surprisingly little is known about changes in the cell wall itself, a challenge for a better understanding of abiotic stress tolerance.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.


We hope you will find this issue insightful and relevant to your practice of glaucoma.In the basic diagnostics section, Sharma et al evaluate the differences between retinal macular thickness and macular volume using optical coherence tomography in primary open angle glaucoma patients and normal subjects. They report a significant difference in the macular parameters as measured on spectral domain-optical coherence tomography (SD-OCT) between the two population subgroups.Secondary angle closure glaucomas are a distinct entity from primary angle closure glaucoma (PACG). The identifiable contributory factors, therefore, present a great diagnostic and therapeutic challenge for the glaucomatologist. Parivadhini et al review the clinical spectrum of this disease subgroup along with management protocols.It is well-known that a proportion of eyes will have residual angle closure, despite a patent peripheral iridotomy. In such patients, a laser peripheral iridoplasty may be a useful treatment to ameliorate appositional angle closure that may be occurring through nonpupillary block mechanisms. Leong et al evaluate, by anterior segment optical coherence tomography, the changes in the anterior chamber angle during the short-term postoperative period after laser peripheral iridoplasty.Lee et al report their initial experiences with the EX-PRESS shunt in a case series of patient with uveitic glaucoma, reporting good intraocular pressure (IOP) control with a propensity for hypotony in the early postoperative period.In an interesting profile of glaucoma patients from Vietnam, Thanh elucidates the number of antiglaucoma medications required to achieve target pressure and correlates the same to severity of disease.There has been considerable interest about the potential use of calcium channel blockers in glaucoma therapy, with both ocular hypotensive effects, and increase in perfusion pressure having been attributed to the drug. Ganekal et al evaluate the effect of 0.125% verapamil and 0.5% diltiazem eye drops on IOP in steroid-induced glaucoma in rabbit eyes and report a considerable drop in IOP, equivalent to that due to topical timolol drops.Kaushik et al report an interesting case of unilateral angle closure glaucoma complicated by an optic disk pit and iridociliary cysts.As always, we look forward to hearing from you.Best wishesShibal BhartiyaTarek ShaarawyTanuj Dada