Langerhans cell histiocytosis (LCH) is a rare inflammatory myeloid neoplasm arising from the proliferation of pathologic Langerhans cells. LCH has a spectrum of presentations predominantly affecting male pediatric patients. As LCH is a relatively uncommon diagnosis, there is no standard of care for treatment of the disease and treatment is based largely on clinical judgment, lesion characteristics, and symptoms at presentation. Here we present a case of unifocal, isolated orbital LCH in a 19‐year‐old young man treated initially with surgical resection. Follow‐up imaging 2 months later demonstrated significant regrowth of the mass and no other sites of disease. The recurrent orbital disease was treated with stereotactic radiosurgery (SRS) to 7 Gy in one fraction. Near complete resolution of the mass was achieved with no recurrence after 1.5 years of follow‐up. SRS for treatment of orbital LCH is a novel treatment not previously described in the literature which may provide benefit in select cases. MRI of the brain demonstrating Langerhans Cell Histiocytosis (LCH) of an orbit. A Birbeck granule is a unique intracellular organelle finding on electron microscopy that is diagnostic for LCH.Langerhans cell histiocytosis is an umbrella term for a group of diseases previously called Histiocytosis X.2The patient is a 19‐year‐old male with a past medical history significant for multiple concussions who presented to a family medicine clinic with complaints of persistently worsening right sided frontal headaches starting 2 weeks prior. At the time, he also noted right eye swelling that did not resolve with cold compress or non‐steroidal anti‐inflammatory medication. The headache pain was described as a pounding sensation with no associated symptoms of photophobia, hyperacusis, nausea, epiphora, fevers, or night sweats. The headaches were initially intermittent but occurred daily until 2 days prior to presentation at which point it became constant and debilitating. The patient also noted intense pressure behind the right eye that was worse when he tilted his head backwards. The patient had no significant contributing family history. The patient's vitals and physical exam were normal including a detailed neurological exam demonstrating no focal deficits. Given the history and physical exam findings, differential diagnoses of migraine, tension headaches, cluster headaches, and infection were felt to be less likely than a possible space occupying lesion in the brain. An MRI of the head . Treatment consisted of 7 Gray (Gy) in one fraction prescribed to the gross tumor volume to cover 95% of the volume at 80% isodose line, as outlined in Figure 3Orbital Langerhans cell histiocytosis (LCH) is a rare condition that primarily affect children and adolescences. Here we present a case of a unifocal orbital LCH lesion in a 19‐year‐old male with successful resolution of recurrent LCH after a single 7Gy Cyberknife stereotactic radiosurgery (SRS). To our knowledge, this case report is the first to describe the use of Cyberknife stereotactic radiosurgery for management of unifocal orbital LCH.Langerhans cell histiocytosis is a proliferation of pathologic Langerhans cells normally found in the body as specialized epidermal dendritic cells. They function to present antigens to other immune cells by traveling through the lymph.Langerhans cell histiocytosis predominantly affects children with an annual incidence of 2 to 10 cases per 1 million children under the age of 15, with 90% of the primary lesions occurring in the head and neck.A diagnosis of LCH can be made through immunohistochemistry or electron microscopy. The presence of langerin (CD207+) or cluster of differentiation 1a (CD1a) on immunohistochemistry or Birbeck granules on electron microscopy are specific for a definitive diagnosis of LCH.Currently there is no standardized treatment for LCH. Treatment approach is individualized based on clinical presentation. A unifocal lesion affecting the skeletal system is the most common form of LCH.If there are doubts about the completeness of the resection or if there is recurrence, radiation at a relatively low dose of approximately 4–8 Gy is suggested for unifocal single system LCH following surgical resection. Radiation is generally more widely used in the adult population as compared to the pediatric population due to possible side effects.Though not generally thought of as a first‐line treatment for LCH, stereotactic radiosurgery (SRS) has been used as first‐line treatment for neurological LCH lesions in several reports that have shown regression and stable tumor outcomes, outlined in Table There are currently nine case reports that present patients who have been treated with either GK or CK SRS at some point in their treatment course. There are seven patients with confirmed CNS LCH lesions who had SRS as part of their treatment course demonstrating resolution and stability of their tumors. None of these patients had SRS as their first or only form of treatment for their lesion. The other two patients treated with SRS had Rosai–Dorfman disease, a histiocyte infiltration into the lymph. Hadjipayanis et al. describes a case of a 52‐year‐old male with lesions in the left petroclival area extending into the cavernous sinus. This was treated with gamma knife SRS with complete resolution at a 13 month follow‐up.For LCH‐specific lesions, Hong et al. reports three adult presentations of acquired diabetes insipidus which was later found to be caused by a posterior pituitary LCH lesion.Emily Etter: Conceptualization; data curation; formal analysis; investigation; methodology; writing – original draft; writing – review and editing. Ben Bosse: Conceptualization; data curation; writing – original draft; writing – review and editing. Yi Peng Wang: Data curation; investigation. Susan Hiniker: Conceptualization; formal analysis; supervision. Justin Oh: Conceptualization; data curation; supervision; writing – review and editing.No funding was received for this study.None of the authors have conflict of interest to declare.Written informed consent was obtained from the patient to publish this report in accordance with the journal's patient consent policy.


Brassica rapa at three different UV radiation intensities  and under two exposure duration regimes. We removed petals from flowers periodically during the flowering period and measured the proportion of the petal that absorbed UV. UV‐absorbing areas increased when plants were exposed to longer periods of UV radiation, and at high UV radiation intensities. UV‐absorbing area of petals of the UV intensity treatments decreased over time in long exposure plants. This study demonstrates that flowers can potentially acclimate to different UV radiation intensities and duration of exposure through an increase in UV‐absorbing areas even after a relatively short exposure time to UV. Such a rapid plastic response may be especially beneficial for dynamically changing UV conditions and in response to climate change.Solar ultraviolet (UV) radiation is known to have significant effects on the development and performance of plants, including flowers. In multiple species, UV‐absorbing floral patterns are associated with environmental conditions such as the solar UV exposure they typically receive. However, it is not known whether plants can increase the UV‐absorbing areas found on petals plastically when in a high‐UV environment. We grew  Flowers can potentially acclimate to different ultraviolet (UV) radiation intensities and duration of exposure through an increase in UV‐absorbing areas even after a relatively short exposure time. It has been extensively cultivated, and is now distributed globally, and found on waste ground, roadside, and both river and stream bankside habitats. To the human eye, the flowers have four bright yellow petals which are arranged symmetrically, with six stamens in the center. In addition, B. rapa has a clear UV‐bullseye , producing up to 20–25 flowers per plant.Field Mustard and Hill  via seve2.2B. rapa plants were exposed to one of two periods of UV exposure prior to anthesis: (i) long‐term UV exposure (~24 days) and (ii) short‐term (~6 days) UV exposure  in a glass greenhouse for 18 days prior to placing them in the chambers for 57 days  with an open top that permitted air flow was placed in the top shelf of both chambers , and Tukey multiple pairwise comparisons indicated that all three UV treatments significantly differed from one another .Three UV radiation intensities were administered across two plant chambers . The natural curvature of d Ashman  indicateA total of 726 pressed flowers were photographed in UV‐A light. All photographs were taken with a Canon EOS 400D camera, fitted with a Baader UBVRI 1¼′′ Photometric U‐filter. The filter permits the transmission of UV‐A light between 320 and 390 nm, peaking with 85% transmission at 350 nm, while removing infrared and daylight wavelengths. The availability of naturally occurring UV light in the daylight spectrum can be limited indoors; the image field was therefore illuminated with a UV‐A torch (Convoy S2+ Nichia UV waterproof LED flashlight) with a peak UV emission at 365 nm. The camera was mounted on a tripod and arm to achieve an overhead view of the pressed specimen. All photographs included a standard scale  and total petal area (mm2) could be accurately measured for a total number of 1871 petals from 644 different flowers from 70 plants, which could be photographed successfully after pressing the petals without damage and with clear focus of the petal  and exposure length (short and long), using a two‐way Kruskal–Wallis test.proportion and petal area, which were modeled using a linear mixed‐effects model (LMM), with individual plant included as a random effect, with flower as a random effect nested within individual plant. We first tested the relationship between radiation intensity treatment , exposure length (short and long), and their interaction on UVproportion and petal area. UVproportion was logit‐transformed using the car package  and UV intensity treatment  on total flower numbers, without a significant interaction between these two factors , with control and low UV intensity plants flowering longer than high UV intensity plants .There was a significant effect of both duration of exposure  and low intensity  plants, with no significant difference between control and low intensity plants . Overall, the UVproportion between them was greater in long exposure plants compared to short exposure plants .For petal areas, there was a significant effect of exposure, but not UV intensity treatment Table ; the pet3.3proportion, there were significant two‐way interactions between plant age and exposure and plant age and UV intensity treatment  can potentially reduce reflected UV damaging pollen; larger bullseyes in artificial flowers provide enhanced protection to UV sensitive pollen, as compared with flowers with smaller UV bullseyes  may be important for the production of flavonoids that are responsible for the UV‐absorbing pigments in floral tissues. While UV exposure early in plant development may increase flavonoid accumulation in both vegetative tissues of B. rapa, and later in floral tissues, it is at present unclear by what mechanism exposure of vegetative tissues to UV affects later floral development (Musil & Wand, We also found that longer exposure increased overall UVproportion, UV intensity also affected petal size in our study. Overall petal area was smallest in long exposure treatments, while no significant difference was observed between UV intensity. In comparison, previous studies have reported a range of negative to positive effects or no changes in petal area in response to UV‐B (see Llorens et al., Besides affecting UVproportion was found to decline as plants aged in the long UV exposure treatment. This effect was not found in short UV exposure treatment. For many plants, there is a decrease in resource allocation and reproductive investment over the flowering period that leads to smaller floral structures, and fewer gametes, and seeds (Ashman & Hitchens, proportion suggests some sort of costs of general reproduction. The fact there is no change in short exposure plants suggests that UV intensity may be an additive cost.Across low and high UV treatments, the UVWhen looking at the effect of plant age on petal area, significant effects were only detected across exposure length and intensity treatment. Controls appeared to show no change over time, whereas long exposure plants seem to show an increase in petal area over time, and in the short exposure group there was no change over time or slight decrease. It is not clear why this was the case, as declines in flower size are normal with plant age in related species (e.g., Williams & Conner, 5proportion in these flowers. We identified a significant positive correlation between the intensity and duration of UV exposure and UVproportion in B. rapa. Even though not tested specifically here, these responses may be important for the hypothesized reduction in exposure to UV radiation of the floral reproductive structures caused by a reduction in the proportion of reflected UV radiation that they receive as a consequence of the increased UV bullseye. Moreover, our study demonstrates that flowers can potentially acclimate to different UV radiation intensities and duration of exposure through an increase in UV‐absorbing areas. Such a rapid plastic response may be especially beneficial in regard to climate change, even though field studies involving multiple species and environmental factors should be employed to fully understand the complexity of UV response in plants.The plastic responses of floral UV‐absorbing area demonstrated in this study indicate the potential for plants to persist when faced with environmental UV‐B change by decreasing flower production but enhancing petal UVThe authors declare no conflict of interests.Appendix S1Click here for additional data file.


This paper re‐examines a well‐established hypothesis postulating that life expectancy augments incentives for human capital accumulation, leading to global income differences. A major distinguishing feature of the current study is to estimate heterogeneous panel data models under a common factor framework, which explicitly accounts for parameter heterogeneity, unobserved common factors (UCFs), and variables' non‐stationarity. In sharp contrast to most previous studies, I find that the impact of health improvements on human capital accumulation turns out to be imprecisely estimated at conventionally accepted levels of statistical significance. I demonstrate that conventional estimates of the educational returns to rising longevity are derived from estimating misspecified models at least partially due to parameter heterogeneity and the presence of UCFs. A major contribution to this line of inquiry postulates that health capital is a key determinant of global income differences. This viewpoint, in particular, is motivated by an early and influential paper by Preston  that docExisting explanations of the positive effect of life expectancy on economic performance primarily rest upon the conventional wisdom that health improvements strengthen human capital investments. The underlying idea is that increased longevity augments motivations to accumulate knowledge by increasing the horizon over which investments in education are discounted Becker, .1 The In addition, several studies provide suggestive evidence of heterogeneity in the impact of health improvements on economic growth and human capital across countries. For example, Cervellati and Sunde  demonstrcommon across the globe. As mentioned previously, the existing literature is suggestive of heterogeneity in the impact of human longevity on education across countries at different stages of the demographic transition and economic development  and a heterogeneous parameter derived from heterogeneous parameter models is central to securing reliable inference on the health‐education relationship ‐averaged panel data models.Identification of the effect of life expectancy on education also requires attention to unobserved common factors (UCFs) that simultaneously affect national health status and human capital accumulation. This concern at least partially stems from the interdependence between world economies. For example, countries could suffer from global common shocks or are subject to the cross‐border diffusion of health technologies and knowledge, thereby affecting a country's health and educational outcomes. However, it is very difficult to identify, measure, and include numerous UCFs in standard regression models. This is because including these common factors in econometric models requires identifying numerous confounding factors, and their unobserved nature makes it inconvenient to measure and identify properly. In addition, incorporating a variety of control variables in the regression may impose substantial constraints on the feasible degrees of freedom. Another concern is that the unknown relationship between UCFs and human capital could result in potential functional form misspecification if they were to be included in the regression. Thus, UCFs generally enter econometric models in the form of disturbance terms in previous studies. Furthermore, earlier studies, to my knowledge, largely ignore the potential non‐stationarity of life expectancy at birth and human capital when investigating the educational returns to health improvements.Motivated by the above concerns, I re‐estimate the effect of life expectancy on human capital accumulation across countries considering the importance of parameter heterogeneity, variables' non‐stationarity, and UCFs. To this end, I construct a balanced panel dataset covering 79 countries over the period from 1964 to 2015. A key distinguishing feature of this study stems from empirical attempts at addressing the above econometric issues. To preview the main findings, I find that life expectancy has a sizable and significant positive effect on human capital accumulation when using conventional panel estimators that do not account for UCFs, variables' non‐stationarity, and parameter heterogeneity. This is in line with most previous studies reviewed earlier. However, the impact of life expectancy on human capital turns out to be imprecisely estimated at conventionally accepted levels of statistical significance once I explicitly account for parameter heterogeneity and UCFs. Conducting several residual‐based diagnostic tests, the preferred estimates provide suggestive evidence that life expectancy, on average, has no statistically significant impact on human capital accumulation. On this basis, I argue that the findings established in the existing literature are derived from estimating misspecified models partly due to the presence of UCFs and parameter heterogeneity.The empirical approach of this study builds upon an emerging body of research revisiting some well‐established hypotheses by applying the common factor framework of Pesaran . For exaThe rest of the paper is organized as follows. Section 22.1http://wdi.worldbank.org/). The dependent variable is the human capital index obtained from the Penn World Table 9.1 . I use annual data from 1964 to 2015 (T=52). Unobserved country‐specific factors are captured by μ. ft stands for a set of UCFs with country‐specific factor loadings τi. β reflects the estimated effect of life expectancy on human capital accumulation. The subscript i on β allows it to differ across cross‐sectional units (parameter heterogeneity).To compare and contrast my findings with conventional estimates in earlier studies, I adopt several panel estimators with different assumptions about data properties, parameter heterogeneity, and the error term. The preferred model is chosen based on some residual‐based diagnostic tests that help identify model misspecification, following Eberhardt et al.  and Eber2.3Conventional panel estimation methods include pooled OLS (POLS) and fixed‐effects (FE) estimators, which have been widely used to explore the health‐education nexus. These estimation techniques require that variables follow stationary processes, the violation of which may yield a spurious relationship among regressors. Although FE models allow the intercept estimates to vary across countries, the estimated slope coefficients, by construction, are identical across cross‐sectional units in the sample . The main conclusion, however, is based on the “second‐generation” panel unit‐root test (CIPS) of Pesaran  because A major concern when exploring the relationship between life expectancy and human capital stems from potential endogeneity bias. This could arise because UCFs may be correlated with LE and HC. Furthermore, reverse causality, if it exists, may also invalidate inference on the education‐health nexus. As such, conventional panel estimators may yield a spurious relationship between longevity and human capital accumulation  and LE. This method is implemented by incorporating cross‐sectional averages for both dependent and independent variables in the regression  and stationary variables. They also require cross‐sectional independence (except for the common correlated effects pooled estimator—CCEP). These estimates are reported for the purpose of comparison. The mean group type estimators, listed in the right column, relax the restrictive assumption of parameter homogeneity. Additionally, the common factor framework (CCE) of Pesaran iI report the estimation results for the dynamic model specification in Table 4.2http://wdi.worldbank.org/). However, the estimated effect of life expectancy on human capital remains insensitive to incorporating additional controls in the regression. It is noteworthy that all the models with additional controls do not pass the CSD test  as an IV for LE. My empirical strategy exploits the fact that health technologies and knowledge transcend national borders and such spillovers decay with cultural distance between countries. Using the genetic distance index of Spolaore and Wacziarg  and (4) of Table 5This paper revisits conventional wisdom in the long‐term comparative development literature asserting that increased life expectancy strengthens incentives for acquiring better human capital skills. This channel of influence is often regarded as a key mechanism through which life expectancy helps shape worldwide income differences. Hence, a wealth of literature has explored the relationship between longevity and education from both theoretical and empirical perspectives. Most empirical studies lend strong evidence to the significant positive effect of health improvements on human capital accumulation. By contrast, Acemoglu and Johnson  empiricaMore specifically, I find that life expectancy has a positive and statistically significant impact on the accumulation of human capital when using conventional panel estimators that assume parameter homogeneity and cross‐sectional independence. However, I argue that these estimates provide an invalid basis for statistical inference because they are derived from estimating misspecified models, evidenced by the serially correlated, non‐stationary, and cross‐sectional dependent residuals of the econometric models. In addition, I adopt the mean‐group type estimators under a common factor framework to control for UCFs and parameter heterogeneity. The heterogeneous parameter estimates reveal that the impact of rising longevity on education is statistically insignificant at conventionally accepted levels when I account for UCFs and parameter heterogeneity. My findings indicate that addressing CSD and parameter heterogeneity is central to securing reliable inference on the relationship between life expectancy and human capital.An important implication derived from my findings is that UCFs and parameter heterogeneity cannot be ignored when estimating the causal effect of life expectancy on human capital accumulation. While incorporating UCFs in econometric models is plagued with various difficulties as argued earlier, the main results suggest that the common factor approach developed by Pesaran  offers aThe author declares that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.Supplementary Material 1Click here for additional data file.


Fibrodysplasia ossificans progressiva is a progressively debilitating condition associated with significant morbidity caused by heterotopic ossification. Recognition of the early signs of hallux valgus and painful soft tissue nodules can assist in the early diagnosis of this condition. Periodic radiographic examination is mandatory to monitor the disease progression.Fibrodysplasia ossificans progressiva is a rare condition with an estimated prevalence of one in two million individuals. The condition is characterized by widespread heterotrophic ossification of skeletal muscles and ligaments. We report the case of an 8‐year‐old female patient and show the radiological progression of the condition. Fibrodysplasia ossificans progressiva (FOP) is a rare condition characterized by widespread heterotrophic ossification and is caused by a mutation in the bone morphogenetic protein (BMP) Type 1 receptor, activin receptor 1A/activin‐like kinase 2 (ACVR1/ALK2).2An 8‐year‐old female patient presented to the University of Pretoria Oral Health centre with progressive limb stiffness and loss of mobility. She was referred due to limited mouth opening and trismus. She was diagnosed with fibrodysplasia ossificans progressiva at 2 years with no other members of the family presenting with similar symptoms. Cone‐beam computed tomography (CBCT) examination revealed ossification of the medial pterygoid muscle on the right Figure  as well Previous radiographic examinations at 2 years of age, revealed calcification of muscles involving the left leg Figure , left anDue to a language barrier, limited communication was possible. No lab records of any blood tests could be traced and the patient was lost to follow‐up.3Fibrodysplasia ossificans progressiva is extremely rare with an estimated prevalence of one in two million individuals.In most cases, the clinical and radiological signs are diagnostic for FOP but genetic testing can be performed. An increased alkaline phosphatase level can also be detected.Treatment guidelines have still not been established as biopsy or surgical intervention often leads to the progression in ossification. Non‐interventional treatment consisting of avoidance of trauma, physiotherapy in combination with non‐steroidal anti‐inflammatory or cyclo‐oxygenase inhibitors during flare‐up episodes, has been advocated.Chané Smit: Conceptualization; investigation; methodology; writing – original draft. André Uys: Conceptualization; investigation; methodology; writing – review and editing.This research did not receive any specific grant from funding agencies in the public, commercial, or not‐for‐profit sectors.The authors declare that they have no conflict of interest.This study was approved by the University of Pretoria, Faculty of Health Sciences Research Ethics Committee (Reference no.: 139/2022). All procedures followed the ethical standards of the Helsinki Declaration of 1975, as revised in 2008.The written patient consent has been signed and collected in accordance with the journal's patient consent policy.


The high‐throughput technologies of RNAi  and CRISPR‐Cas9‐mediated gene knockout have greatly facilitated unbiased discovery of such drug–drug synthetic lethality. Here, we will discuss two examples of drug–drug synthetic lethality, which have been translated from ‘bench to bed’, highlighting the power of high‐throughput screening in guiding the design of clinical combination therapies.3In the early twenty‐first century, RNAi technology was introduced, allowing high‐throughput screening in cancer cells driven by specific oncogenic mutations. A particularly interesting example of the use of this technology relates to the proper therapy selection of different BRAF mutant cancers. Even though activating mutations in the BRAF oncogene (BRAF(V600E)) were found in some 70% of primary melanomas, some 10% of colorectal cancers and some 30%−70% of papillary thyroid carcinoma, clinical responses to the highly selective small‐molecule inhibitor of the BRAF(V600E) oncoprotein, vemurafenib (PLX4032), differed widely, ranging from a response rate of approximately 80% in melanoma to only 5% in BRAF mutant colorectal cancers (CRCs).To investigate the molecular mechanism responsible for the intrinsic resistance of BRAF(V600E) CRCs to vemurafenib, Prahallad et al. performed a short hairpin RNA (shRNA)‐based genetic screen investigate whether inhibition of any of the 518 kinases in the human genome synergizes with BRAF(V600E) inhibition.4RNAi technology is prone to off‐target effects, meaning that RNAi targeting of one gene may accidentally co‐target another gene. Luckily, not long after RNA interference, the CRISPR gene editing technology was discovered, which can perfectly overcome the off target issues of RNAi technology and improve efficiency and accuracy.In 2021, our group discovered a novel rational combination for the treatment of liver cancer in preclinical investigations. The pre‐clinical data were supported by encouraging evidence of clinical benefit in a proof‐of‐concept trial. The multi‐kinase inhibitor lenvatinib has been approved by the FDA for the first‐line treatment of unresectable hepatocellular carcinoma (HCC), but the overall response rate is only about 24%.5Both studies emphasize the advantage of using functional genetic screens to discover novel and effective drug–drug synthetic lethality interactions to improve the efficacy of targeted therapies (See Figure Furthermore, the use of two drugs may still not be enough. Despite combinatorial targeted therapies based on the latest understanding of signaling circuitry,Using combination therapies to treat cancer may seem simple in theory, but difficult in practice because of the many possible combinations of the present arsenal of cancer drugs. Rational drug combinations that show strong synergy based on insights in how signaling pathways communicate are a logical path forward.The National Natural Science Foundation of China Grant Numbers: NSFC82222047 and NSFC82073039; The Program of Shanghai Academic/Technology Research Leader, Grant Number: 22XD1423100.The authors declare no conflict of interest.


Protective defense mechanisms are well documented across the animal kingdom, but there are still examples of antipredator defenses that do not fit easily into the current conceptualization. They either fall within the intersection of multiple mechanisms or fail to fall neatly into pre‐existing categories. Here, using Endler's predatory sequence as a framework, we identify problematic examples of antipredator defenses, separating them into protective mechanisms that are difficult to classify and those which act sequentially depending on context. We then discuss three ways of improving underlying terminological and definitional problems: (1) issues with English and polysemy, (2) overlapping aspects of similar mechanisms, and (3) unclear definitions. By scrutinizing the literature, we disentangle several opaque areas in the study of protective defense mechanisms and highlight questions that require further research. An unclear conceptual framework for protective defense mechanisms can lead to misconceptions in understanding the costs and benefits of defenses displayed by animals, while interchangeable terminologies and ambiguous definitions can hinder communication in antipredator studies. Although the main forms of antipredator defenses in animals are well defined, there are still many examples of overlapping mechanisms and others that do not fall neatly into pre‐existing categories. In our review, we systematically discuss these problems and suggest ways of improving underlying terminological and definitional problems. By scrutinizing hitherto unexplored examples of confusing antipredator defenses across taxa, we disentangle opaque areas in the study of protective defense mechanisms and highlight questions that require further research. They concluded that the behavior functioned to deflect predatory attacks to the torso. They further investigated the phylogenetic relationships among four lizard families  and found that the deflection hypothesis is supported in all four of them, but only two families, Iguanidae and Gekkonidae, provided evidence supporting the pursuit deterrence hypothesis. Thus, it is likely that the deflection hypothesis is the ancestral function, and that pursuit deterrence is the less common derived function.The study by Telemeco et al.  was the C. collaris; Figure Leiocephalus spp.) cannot distinguish between pursuit deterrence or deflection when approached by humans because their findings support both hypotheses  and keeled box turtle (C. mouhotii) have yellow stripe(s) on their dark brown carapaces, for example  exposed, then these stripes will likely match the leaf litter background  is a brightly colored toxic caterpillar that has alternating orange‐and‐black horizontal stripes on its body  using model prey  exists in two forms, with the red form mimicking the seven‐spot Coccinella septempunctata and the melanic form mimicking the four‐spot Exochomus sp.  have shown that the red two‐spot A. bipunctata (mimic) is only mildly toxic compared to the highly toxic seven‐spot C. septempunctata (model), confirming the quasi‐Batesian nature of the mimicry  leading to misclassification. This section identifies three exemplar polysemies and suggests alternative words to use.3.1.1Colloquially, as well as more generally in behavioral biology, flash behavior could mean the sudden or intermittent exhibition of coloration. Indeed, flash as a verb simply describes an intermittent display of coloration by a prey  uses a flashing coloration at the rear of her abdomen to attract males rather than a way of responding to predators  flashes iridescent blue coloration when disturbed which has been termed a “flash mechanism”  emits bioluminescent flashes when predatory seals are present  is achieved through light reflection and acts to reduce the chance of a successful attack by the predator on any one individual  incorporate egg sacs or detritus into silk decoration on their webs. For example, the orb spider (Cyclosa mulmeinensis) uses its prey pellet or egg sac to decorate its web which may increase its survival chances  reveals its conspicuous black and white underwings in its defensive posture when it perceives a threat which is considered as deimatic behavior  during flight is considered flash behavior  exhibits a catalepsy phase 10 times longer than the aposematic gregarious morphs and is thought to enhance camouflage efficacy in the solitary state. This catalepsy should, therefore, be considered as freezing, not tonic immobility since it operates during the detection phase of the predation sequence ”  have been described as showing death/illness feigning and attacking small scavenging fish when they lower their guard  flew with a minimum positional angle and wingbeat frequency more similar to the unpalatable Pachliopta aristolochiae (model) than its non‐mimetic form (the cyrus form). Similarly, an ant‐like resemblance is seen in over 13 spider families, including mimicry in both morphology and behavior  which lack chemical defense traits mimic the walking behavior of chemically‐defended ants (Formica sp.) and also demonstrate their characteristically short pauses  bear a highly similar color pattern as the evasive flesh fly suggesting Batesian evasive mimicry underlies the resemblance  mimicry encompasses mimicking the visual appearance of the hard‐to‐catch model, whereas locomotor mimicry focuses on mimicking the typical escape behavior of an unpalatable model. Unprofitability in evasive mimicry arises from the difficulty of catching the prey, whereas the unprofitability in locomotor mimicry comes from the distastefulness of the model.In summary, this section explores overlapping aspects of different protective coloration mechanisms while highlighting their distinctions. To reduce confusion in terminology, we emphasize the need for precise definitions and accurate categorization.3.3There are unclear definitions of protective defense mechanisms that need attention. Here, we try to identify some grey areas.3.3.1Pursuit deterrence has been defined as “signals used by prey that apparently convince predators not to pursue them” Caro, . It can Otosphermophilus beecheyi) display tail‐flagging behavior that is both related to their alertness and the likelihood of a snake's presence  shown toward foxes (Vulpes vulpes)  were chased by merlins  for longer periods of time and were more likely to be caught compared to skylarks that sang well. Although Cresswell did not directly test the energy cost of singing in skylarks, other studies have shown that singing during flight is energetically demanding in both skylarks and other birds  serves as a perception advertisement toward ambushing predators  are less likely to chase after gazelles that stot.There are other more challenging examples of pursuit deterrence which are harder to classify into either perception or quality advertisement. For example, many ungulates stot toward the predator, and stotting in Thomson's gazelles  is a species in the Sacoglossa group that resembles the Bryopsis toxic algae other striking algal mimic in Sacoglossa” and inferred it to be an example of Batesian or Müllerian mimicry. However, it is possible to consider C. mediterranea as a masquerader of the algae, similar to a twig‐resembling caterpillar or leaf‐resembling butterfly  is an example of olfactory crypsis , then masquerade actually overlaps conceptually with Batesian mimicry. This is because the living model, such as a leaf, may receive an increase in attack rate from birds  due to the presence of the leaf‐resembling comma butterfly (the masquerader)  Deimatic and flash behavior both involve sudden displays of conspicuous color patches but differ in their effect on the predator. (2) Tonic immobility, freezing, and some forms of aggressive mimicry involve stopping all movements but are performed at different predation stages to achieve distinct purposes. (3) Evasive and locomotor mimicry both involve the movement of the model, but the former involves mimicking the appearance of the hard‐to‐catch model, whereas the latter focuses on mimicking the escape behavior of an unpalatable model. Although significant progress has been made in clarifying these mechanisms to avoid misconceptions with other terms, there are still studies that refer to these pairs of mechanisms interchangeably.Finally, there are unclear definitions that need addressing. Two sets of mechanisms, pursuit deterrence and masquerade‐Batesian mimicry, require definitions to be reformulated. The definition of pursuit deterrence may need to recognize vigilance, perception and quality advertisement as a gradation of increasing cost Table  and futuIn conclusion, the study of animal antipredator defense mechanisms still demands more precise terminology and clearer definitions given its complexity. To avoid potential confusion, the names of new mechanisms should be carefully crafted to avoid polysemic words that may be applied in other contexts unrelated to defense mechanisms. Clear definitions are also essential to guarantee precision in interpreting experimental findings and definitions should be framed in a manner that is inclusive of all currently known putative examples of defense mechanisms across different taxa.It is also vital to acknowledge that animals often employ multiple mechanisms that may not be mutually exclusive ; writing – original draft (lead); writing – review and editing . Tim Caro: Conceptualization ; validation (lead); writing – review and editing .The authors declare no conflicts of interest.


Glioblastoma multiforme (GBM) is an aggressive brain tumor with poor prognosis and high mortality, with no curative treatment to date as limited trafficking across the blood–brain barrier (BBB) combined with tumor heterogeneity often leads to therapeutic failure. Although modern medicine poses a wide range of drugs that are otherwise efficacious in treating other tumors, they often do not achieve therapeutic concentrations in the brain, hence driving the need for more effective drug delivery strategies. Nanotechnology, an interdisciplinary field, has been gaining immense popularity in recent years for remarkable advancements such as nanoparticle (NP) drug carriers, which possess extraordinary versatility in modifying surface coatings to home in on target cells, including those beyond the BBB. In this review, we will be highlighting recent developments in biomimetic NPs in GBM therapy and how these allowed us to overcome the physiological and anatomical challenges that have long plagued GBM treatment. Astrocytes are the most prevalent cells in the brain, outnumbering neurons by more than five‐fold and serving an equally vital function. Beyond providing nutrients to neurons, they play key roles in regulating the blood–brain barrier (BBB) and maintaining interstitial fluid homeostasis—a crucial factor to facilitate synaptic transmission.The BBB consists of astrocytes anchoring pericytes and extracellular matrices to the basement membrane of an endothelial cell layer that forms tight junctions (TJs), which are responsible for regulating the movement of substances across the BBB. The diffusion of small lipophilic molecules is facilitated by important transmembrane proteins within TJs, which include zona occludens (ZO), claudins, and junctional adhesion molecules.This is not necessarily the case with GBM however, as the rerouting of vasculature to the growing tumor often destabilizes the BBB. The “leaky barrier,” or blood–brain tumor barrier (BBTB), allows macromolecules designed for targeted drug delivery to bypass the normally impregnable BBB. Unfortunately, high‐grade gliomas rapidly infiltrate surrounding tissues where the BBB is not yet compromised, making the BBTB as a sole delivery route via the enhanced permeability and retention (EPR) effect an unreliable strategy.Nanoparticles range in size from 1 to 100 nm, and can be synthesized using inorganic, polymeric, or organic materials.In this review, we discuss current GBM therapeutic strategies and their associated challenges, as well as demonstrate how NPs could be a novel approach to overcome the limitations in conventional GBM treatment. In Figure 2The current standard of care relies solely on relatively conventional treatments, and patient outcomes remain largely dismal despite decades of development. Surgical cytoreduction of the tumor mass is the current cornerstone of treatment, and aims to alleviate tumor mass effects while collecting samples for immunohistochemical profiling to identify potential targetable markers.MSH2 and MSH6.Temozolomide, a lipophilic molecule that can cross the BBB, is a monofunctional alkylating agent which methylates DNA and creates cytotoxic DNA adducts such as O6‐methylguanine (O6MG). O6MG causes the misincorporation of thymine in place of cytosine during DNA synthesis, resulting in base‐pair mismatches. The mismatch repair (MMR) system will be activated and halt DNA synthesis. If repair is unsuccessful, DNA replication fails and double‐strand breaks will lead to cytotoxic lesions and apoptosis.In addition to TMZ resistance, GBM's angiogenic tendencies also pose a significant hurdle to treatment.Another major factor limiting the efficacy of current GBM treatment is the highly heterogenous nature of the tumor. Even within the same tumor mass, there can be multiple clonal and subclonal populations, each marked by distinct genetic alterations.Novel therapeutic strategies such as vaccine therapy using tumor peptides, tumor cells, and dendritic cells have also been gaining traction in recent years. A clinical trial targeting cytomegalovirus pp65 protein, which is found in a number of GBM tumors, has shown significant increase in PFS as well as OS.Many drugs have been explored for GBM treatment but none have yet to succeed in the clinical setting. While the above‐mentioned examples have shown some efficacy in eliminating GBM tumor cells, the inconsistency in therapeutic outcomes drives the search for new pharmacologic agents. Many of the identified pharmacologic agents do not reach therapeutic concentrations in the brain due to the highly impermeable BBB.33.1One way to bypass the BBB is by directly administering drugs into the interstitium of the brain, hence minimizing systemic side effects. An accepted approach is by encapsulation within a bioresorbable implant which is deposited into the resection cavity following tumor resection.Another technique for intracranial administration of drugs is convection‐enhanced delivery (CED), which utilizes a catheter inserted directly into the tumor to generate a pressure gradient that disseminates the drug to the target site. This enables delivery of substantial drug volumes and allows therapeutic concentrations to be achieved in the brain. However, as with GLIADEL wafers, they have limited efficacy in targeting infiltrative GBM tumor cells at distant sites. The invasive nature of this technique also decreases its attractiveness to both patients and clinicians. It is hence unsurprising that no drugs have yet been approved for use with CED despite multiple clinical trials since its inception in 1994.3.2An alternative strategy being explored to overcome the impermeable BBB is by inducing a transient opening to grant passage for therapeutics delivery. MRgFUS is a noninvasive method that combines a low intensity FUS together with systemically injected microbubbles to mechanically disrupt the BBB without causing heat injury.3.3Over the years, many types of NPs have been developed to enhance permeability across the BBTB and concentrate drug delivery at target sites to minimize systemic toxicity.Despite the immense potential of these NPs in treating GBM, their efficacy remains limited by their susceptibility to clearance by the reticuloendothelial system (RES) due to the presence of numerous immunogenic molecules on their surface. This significantly decreases the number of NPs that eventually reach their target.4As with most drugs, the efficacy of traditional NPs is limited by their relative inability to pass through the BBB, with research showing that only 0.7% of the administered NP dose is successfully delivered to tumors in the brain.As illustrated in Figure In this review, biomimetic NPs are classified into four major categories—protein‐based, ligand‐based, cell membrane‐based, and natural transporter‐based types, all of which present unique solutions to the limitations of traditional NPs.4.1Many serum proteins circulating in our bloodstream have been explored as possible drug delivery carriers for GBM treatment. Albumin, the most abundant protein in plasma, has been extensively explored for biomimetic NP design. Its long circulatory half‐life enables albumin‐based NPs to remain in circulation for a longer period of time and accumulate in tumors at higher concentrations.Human H‐ferritin (HFn) NPs, designed by Fan et al., were able to traverse the BBTB and accumulate within GBM tumor cells, while sparing surrounding normal brain tissue.4.2Biomimetic NPs that are coated with natural GBM‐specific ligands such as folate and lactoferrin have been developed in many studies. Folate, also known as vitamin B9, is a commonly used ligand to increase tumor specificity of biomimetic NPs.l‐malic acid) NPs to target endothelial cells in the BBB which overexpress transferrin receptors, allowing these NPs to cross the BBB with greater ease. An additional anti‐EGFR mAb (cetuximab) was also added to enable these NPs to target GBM tumor cells with increased EGFR expression.Monoclonal antibodies (mAb) specific for receptors found on GBM tumor cells can also be synthesized to increase homing capacity of NPs. An antitransferrin receptor mAb has been designed and used to coat poly are often overexpressed in GBM and are responsible for the movement of many molecules across the BBB. An apolipoprotein B derived peptide, ApoB29 mer peptide, is a ligand that binds with high affinity to LDLRs and was used in a study by Seo et al.In another study, albumin‐based NPs were conjugated with a short peptide that can bind to transferrin receptors on the BBB and GBM tumor cells.Many other ligands that have been used to synthesize biomimetic NPs belong to a novel category called cell penetrating peptides (CPPs). CPPs are made up of short polypeptides of about 5–30 amino acids. The iRGD peptide is an example of a CPP which binds to integrins and neuropilin‐1 receptors to allow endocytosis of substances into cells. It is often used for targeting angiogenic endothelial cells in GBM. In a study conducted by Liu et al., NPs with iRGD CPP demonstrated increased uptake and were able to selectively accumulate in GBM cells at higher concentrations in preclinical experiments.4.3The key advantage of NPs coated with natural cell membranes is their hypoimmunogenic physicochemical properties as they are able to evade immune surveillance by the RES, thereby increasing their time in circulation.4.3.1Among the various types of immune cells, macrophage‐coated NPs have been used in several experimental studies for GBM therapy for their exceptional ability in traversing the BBTB. Macrophage‐coated near‐infrared Ib (NIR‐Ib) fluorescent dye IR‐792 NPs (MDINPs) used with NIR‐Ib imaging‐guided photothermal therapy in an orthotopic mouse model has shown heartening results in treating GBM.In addition, NK cell membranes have also been used as coating substances for biomimetic NPs in GBM therapy. NK cell nanorobots with aggregation‐induced emission (AIE) characteristics (NK@AIEdots) have been developed by coating NK cell membranes on an AIE‐active conjugated polymer.4.3.2Cancer cells have also been explored as potential sources of cell membranes to synthesize membrane‐coated NPs. The tumor specific proteins used to coat these NPs include cell adhesion molecules such as cadherins, selectins, and integrins, which enable homologous binding to the source cell line to bestow a potent homing effect.4.4+ T cell‐mediated immunity and enabling the development of long‐term immunological memory against subsequent tumor antigens from recurrent GBM.Naturally occurring transporters in the body such as high‐density lipoprotein (HDL), low‐density lipoprotein (LDL) and exosomes can be used as the base for biomimetic NP design. HDLs, in particular, are a popular choice for nanocarriers, as they are able to remain in circulation for an extended period of time, allowing accumulation of NPs at tumor sites. Similar to natural cell membranes used to coat biomimetic NPs, HDLs already possess in‐built ligands such as apolipoprotein A1 which recognize HDL scavenger receptors in the BBB and on GBM tumor cells.Low‐density lipoproteins are structurally similar to that of HDLs and have apolipoproteins that are recognized by in vis found on endothelial cells lining the BBTB and GBM tumor cells. In a study, synthetic nano‐LDL particles were synthesized by combining a peptide containing a lipid binding motif and the LDLR binding domain of apolipoprotein B‐100 with a lipid emulsion.Exosomes, a type of extracellular vesicle, are naturally occurring transporters responsible for the intercellular transfer of molecules.Other natural transporters that have been explored for drug delivery to GBM tumor cells are viruses. Viral vectors have long been used as a delivery platform for gene therapy. Retrovirus (RV), adenovirus (ADV), adeno‐associated virus (AADV), and herpes simplex virus‐1 (HSV) are the most commonly used viral vectors for GBM gene therapy.Recent developments in the field have given rise to viral‐mimetics that can be conjugated with NPs to improve homing capacity to the CNS. Gold‐liposome NPs conjugated with brain targeting peptides—rabies virus glycoprotein (RVG), and apolipoprotein E, were designed to improve the delivery of RNA interference (RNAi) for GBM treatment.The numerous examples listed above have highlighted improved antitumoral outcomes with biomimetic NPs when compared to traditional NPs. However, despite all these promising developments in the field of biomimetic NPs, translation from preclinical to clinical setting remains a work in progress. Henceforth, more effective targeting strategies that augment current biomimetic NPs are imperative.5As mentioned previously, efficacy of most drugs including NPs, is often reduced due to inadequate BBTB penetration and the tricky characteristics of solid tumors, especially tumor heterogeneity. Current GBM treatment is also limited by the immunosuppressive TME, irregular vasculature causing an uneven distribution of systemically administered drugs,5.1Tumor specificity is a key, yet often limiting factor in the efficacy of biomimetic NPs, especially in a highly heterogeneous tumor like GBM. One method to resolve this issue is the use of chimeric antigen receptors (CARs). CARs consist of an extracellular antigen‐specific domain comprising monoclonal antibody fragments that is fused to an intracellular domain made up of the CD3ζ chain of the TCR and costimulatory molecules for signaling purposes.These molecules have all been clinically verified as safe targets of CAR‐T therapy for GBM, but only two CAR‐T therapies specific to CD19+ B cell malignancies are currently approved by the FDA.That being said, the combination of CAR‐T cell technology and biomimetic NPs to create a new therapeutic vehicle could potentially overcome the immunosuppressive TME in GBM. As a single CAR‐T cell membrane can be modified to carry a tailored blend of tumor specific antigens, they are especially useful in targeting heterogeneous tumors like GBM.5.2Besides CAR‐T cell membranes, GBM tumor cell membranes can also be used for homologous targeting of NPs to GBM. Additional TME‐responsive elements can then be used to further increase the efficacy of tumor killing. Recently, a team of researchers formulated GBM cell membrane‐coated NPs, which demonstrated superior BBB penetration and GBM targeting through homotypic recognition, in comparison to bare NPs. These NPs contain lactate oxidase which convert lactic acid present in high concentrations in the TME into pyruvic acid and hydrogen peroxide. Pyruvic acid suppresses tumor cell growth while hydrogen peroxide reacts with the delivered bis oxalate to release energy, which is subsequently utilized by the photosensitiser chlorin e6 for the generation of singlet oxygen to eliminate GBM cells.Another example would be the creation of GBM cell membrane‐coated NPs enveloping a pH degradable acetal grated dextran inner core loaded with TMZ and cisplatin.5.3Another strategy to increase target specificity of biomimetic NPs is the usage of aptamers, which are single‐stranded oligonucleotides that possess different configurations and bind to target molecules such as membrane‐bound proteins.5.4A large proportion of current research aims to enhance specificity of drug‐delivery carriers for the BBTB and GBM tumor cells. However, studies on the mechanisms to increase drug uptake from the interstitial space into tumor cells are far and few between. As mentioned earlier, macropinocytosis is a form of endocytosis that allows the nonselective uptake of proteins and extracellular fluids into cells via vesicles called macropinosomes.5.5After extravasating from blood capillaries in the BBB, NPs enter an interstitial space containing interstitial fluid that bathes the parenchymal cells of the brain and spinal cord.Besides capitalizing on ligand‐receptor interactions to improve NP uptake into GBM cells, the transport of NPs through the interstitial space can also be enhanced by breaking down the dense ECM using ECM‐digesting enzymes such as collagenase and hyaluronidase.Human recombinant hyaluronidase has also been successfully added to red blood cell membrane‐coated NPs and this conjugation was observed to enhance NP diffusion in the hyaluronic acid‐rich matrix surrounding PC3 prostate cancer cells.5.6The combination of biomimetic NPs with other specific types of therapy can also selectively enhance different therapeutic effects. PLGA nanovectors encapsulating silver NPs were coated with chlorotoxin (CTX) to target matrix metalloproteinase‐2 (MMP‐2) and chloride channel‐3 (CIC‐3), both of which are highly expressed on GBM cells.6Despite significant progress in the treatment of other cancers over the years, current treatment strategies for GBM often fall short of treatment goals. The current array of therapeutic tools remains inadequate, and is often blunted by acquired drug resistance by rapidly evolving GBM cells. To effectively pioneer a new paradigm of GBM treatment, overcoming the major obstacles faced today is paramount. One of which is the impermeable BBB, which precludes the penetration of several therapeutic drugs into the CNS. The need for improved drug delivery methods across the BBB so that better therapeutic strategies can be devised is clear, and biomimetic NPs are an extremely attractive proposition, especially if combined with other strategies such as CAR‐T technology and ECM‐digesting enzymes. The increased tumor specificity and decreased immunogenicity conferred by these particles could usher in a new age of improved therapeutic outcomes. Although further studies and enhancements are necessary to optimize these NPs for clinical use, nanotechnology is undoubtedly a field that possesses a wealth of untapped potential that could radicalize the treatment of GBM.Xin Yuan Lim: Conceptualization ; writing – original draft ; writing – review and editing . Sharah Mae Capinpin: Writing – original draft ; writing – review and editing . Nagarjun Bolem: Supervision (supporting); writing – original draft (supporting); writing – review and editing (supporting). Aaron Song Chuan Foo: Supervision (supporting); writing – original draft (supporting); writing – review and editing (supporting). Wai‐Cheong George Yip: Supervision (supporting); writing – original draft (supporting); writing – review and editing (supporting). Daniel Boon Loong Teh: Conceptualization (lead); writing – original draft (lead); writing – review and editing (lead).The authors declare no conflicts of interest.https://publons.com/publon/10.1002/btm2.10483.The peer review history for this article is available at


Folsomia candida, we investigate the impact of corridor quality on metapopulation persistence under a range of simulated droughts, a key stressor for this species. We manipulated both drought severity and the number of patches affected by drought across landscapes connected by either good‐ or poor‐quality corridors. We measured the time of metapopulation extinction, the maximum rate of metapopulation decline, and the variability of abundance among patches as criteria to evaluate the persistence ability of metapopulations. We show that while drought severity negatively influenced the time of metapopulation extinction and the increase in drought patches caused metapopulation decline, these results were mitigated by good‐quality corridors, which increased metapopulation persistence time and decreased both how fast metapopulations declined and the interpatch variability in abundances. Our results suggest that enhancing corridor quality can increase the persistence of metapopulations, increasing the time available for conservation actions to take effect, and/or for species to adapt or move in the face of continued stress. Given that fragmentation increases the isolation of habitats, improving the quality of habitat corridors may provide a useful strategy to enhance the resistance of spatially structured populations.Corridors with good‐quality habitats maintain the spatial dynamics of metapopulations by promoting dispersal between habitat patches, potentially buffering populations, and communities against continued global change. However, this function is threatened by habitats becoming increasingly fragmented, and habitat matrices becoming increasingly inhospitable, potentially reducing the resilience and persistence of populations. Yet, we lack a clear understanding of how reduced corridor quality interacts with rates of environmental change to destabilize populations. Using laboratory microcosms containing metapopulations of the Collembola  Corridors maintain the spatial dynamics of metapopulations by promoting dispersal between habitat patches, potentially buffering populations and communities against continued global change. Using a lab microcosm experiment, we demonstrate how corridor quality maintains the persistence of metapopulations in the face of extreme climate change. Drought is a known abiotic stressor to F. candida populations, as they need a high moisture to survive and reproduce. By monitoring the changes in metapopulation abundance in microcosms for a relatively long‐period , we can measure the effects of corridor quality interacting with drought severity and increase in drought patches at the time of extinction, the maximum rate of metapopulation decline, and the variability of abundance among patches.Here, we investigate the effect of corridor quality on the persistence of metapopulations under drought stressors. We manipulated both drought severity and the number of patches affected by drought in landscapes connected by either good‐ or poor‐quality corridors. We use 3D‐printed four‐patch and four‐corridor microcosms containing the soil Collembola 22.1Folsomia candida . F. candida was reared in Petri dishes at room temperature (c. 19°C), with dry yeast provided as a food resource. Although F. candida can survive in drought conditions for short periods by temporarily changing their physiological response  connected by four corridors (c. 28 cm in length and 1 cm in width) in a square (2 × 2) configuration, with each habitat patch connected by two corridors. Both habitat patches and corridors were filled with a mixture of plaster of Paris and black dye (c. 0.5 cm in thickness). Plaster of Paris acts as a substrate for the Collembola to live on which keeps moisture, while the black dye makes Collembola more visible for counting  and consisted of four habitats patches (circular: n = 30) throughout the experiment, and thus food was not a limiting factor to growth. Collembola were reared in the microcosms for 3 weeks prior to the start of treatments to allow metapopulations to stabilize, with 2 mL water and 7 mg of food added each week to each patch. Clear lids were used to cover the microcosms and stop them drying out too quickly, and all microcosms were placed in dark rearing room to minimize light exposure.As Collembola are delicate and not easy to handle, they were inoculated into the four patches of each landscape by tapping individuals (mean ± SD = 423 ± 317) into each microcosm (week 1 of the experiment). During the experimental setup, a potential confounding effect was accidentally introduced whereby microcosms were not randomly assigned to treatment after introducing the Collembola. This may have caused systemic differences in population‐level responses to treatments  with a zero‐inflated negative binomial distribution to investigate how corridor quality, drought severity, and increase in drought patches influence the variability of abundance among patches through time. The variability of abundance among patches was defined as the standard deviation of abundances between the four habitat patches. Microcosm identity was included as a random effect to account for the potential differences between microcosms including their starting densities. We included time, corridor quality, drought severity, increase in drought patches, and their interactions as fixed effects. Of particular interest was how experimental factors  and time interactively affect the changes in variability, thus we also include their two‐way interactions in the fixed effect. Time and experimental factors  were included as a zero‐inflation parameter to account for the overdispersion of zero counts after extinction occurred. Data collected before drought treatment started  was excluded from model fitting. The model was fitted using “glmmTMB” package )) where Y was total abundance and X was time; d was the high asymptote; b was the maximum slope; e was the time at the maximum slope, were all estimated by models. This is a three‐parameter logistic function, and we chose this model due to its simplicity and utility  for each microcosm, we used a GLM with a Gaussian distribution to fit the log‐transformed data, where corridor quality, drought severity, increase in drought patches, and their two‐way interactions were included as explanatory variables.Finally, we used nonlinear regression curves with a three‐parameter logistic function to fit the changes in total population abundance in each habitat network over time. Microcosms either in good‐ or poor‐quality corridor treatment experienced the same rearing conditions prior to the start of the drought treatments , thus we only fit the nonlinear regression curves with the data on metapopulation abundance from week 4 to week 16 Appendix . The mods Figure . Again, 3R2 = .527) showed that drought severity negatively affected the time of metapopulation extinction  showed that time negatively affected the variability of patch abundance  showed that corridor quality negatively affected the maximum rate of metapopulation decline, with their declining rate being slower in good than poor corridor quality treatment ; formal analysis ; funding acquisition ; investigation ; methodology ; writing – original draft . Jane Memmott: Supervision . Christopher F. Clements: Supervision .The authors declare no conflict of interests.Appendix S1Click here for additional data file.


Sir,et al.,[I read with interest the recent report on “Tuberculin skin test for the diagnosis of latent tuberculosis during renal replacement therapy in an endemic area: A single center study” by Agarwal et al., and haveHost resistance to infection is primarily mediated by cellular immunity which is deficient in patients with chronic kidney disease (CKD). The occurrence of infections including tuberculosis (TB) is therefore high in such patients. In addition, various studies have reported the incidence of TB in patients with CKD and on maintenance hemodialysis to be 6–16 times that of general populations.1 Impaired


Primary‐prevention implantable cardioverter‐defibrillators (ICDs) reduce total mortality in patients with severe left ventricular systolic function. However, only a minority of patients benefit from these devices. We designed the Prospective Observational Study of Implantable Cardioverter‐Defibrillators (PROSE‐ICD) to identify risk factors and enhance our understanding of the biological mechanisms that predispose to arrhythmic death in patients undergoing ICD implantation for primary prevention of sudden death.This is a multicenter prospective cohort study with a target enrollment of 1200 patients. The primary end point is ICD shocks for adjudicated ventricular tachyarrhythmias. The secondary end point is total mortality. All patients undergo a comprehensive evaluation including history and physical examination, signal‐averaged electrocardiograms, and blood sampling for genomic, proteomic, and metabolomic analyses. Patients are evaluated every 6 months and after every known ICD shock for additional electrocardiographic and blood sampling. As of December 2011, a total of 1177 patients have been enrolled with more nonwhite and female patients compared to previous randomized trials. A total of 143 patients have reached the primary end point, whereas a total of 260 patients died over an average follow‐up of 59 months. The PROSE‐ICD study represents a real‐world cohort of individuals with systolic heart failure receiving primary‐prevention ICDs.Extensive electrophysiological and structural phenotyping as well as the availability of serial DNA and serum samples will be important resources for evaluating novel metrics for risk stratification and identifying patients at risk for arrhythmic sudden death.http://clinicaltrials.gov/ Unique Identifier: NCT00733590.URL:  Unfortunately, this metric is neither highly sensitive nor specific and does not take into consideration other genetic or electrophysiological determinants for arrhythmogenesis. As a result, there is substantial interest in identifying more reliable predictors that could help to discriminate which patients are most likely to benefit.Sudden cardiac death (SCD) is the most common cause of death in the United States, affecting >300 000 individuals yearly.The Prospective Observational Study of Implantable Cardioverter‐Defibrillators (PROSE‐ICD) is a multicenter prospective observational study of patients undergoing ICD or cardiac resynchronization therapy (CRT) implantation for primary prevention of SCD. Its aim is to identify risk factors and enhance our understanding of the biological mechanisms that predispose to arrhythmic SCD, with the hope of developing a paradigm to identify individuals who stand to benefit the most and the least from primary prevention ICDs. Patients undergo extensive phenotyping and biobanking of DNA and serum at the time of implantation, at 6‐month intervals throughout follow‐up, and after ICD shock events. This article presents an overview of the PROSE‐ICD design, a descriptive analysis of the demographics of the study participants, and anticipated event rates and enrollment plans.clinicaltrials.gov identifier NCT00733590) of patients undergoing ICD implantation for primary prevention of SCD on the basis of criteria outlined in current practice guidelines.12 The study is being conducted at 4 centers in the United States: the Johns Hopkins Hospital and Bayview Medical Center, Baltimore, MD; the University of Maryland Hospital, Baltimore, MD; the Washington Hospital Center, Washington, DC; and the Virginia Commonwealth University Hospital, Richmond, VA.PROSE‐ICD is a multicenter prospective cohort study  ischemic cardiomyopathy  with an ejection fraction of ≤30% and (2) ischemic or nonischemic cardiomyopathy with an ejection fraction ≤35% and stable heart failure symptoms (New York Heart Association [NYHA] Class II to III) on optimal pharmacotherapy . In addiAt enrollment and prior to ICD implantation, each patient undergoes a comprehensive history and cardiovascular physical examination, electrocardiographic (ECG) evaluation, cardiac imaging to assess ejection fraction, and a blood draw. ECGs are recorded digitally  and include a standard 12‐lead recording as well as a high‐resolution 5‐minute rhythm strip using 3 orthogonal leads. Adjudicated automated readings of the 12‐lead ECGs are performed for rhythm determination and conduction abnormalities. The rhythm strip is analyzed by computer‐based algorithms for signal averaging and for measuring heart rate and QT variability parameters. Heart rate and QT variability analyses are performed in the time and frequency domains after excluding ECGs exhibiting frequent atrial or ventricular ectopy, atrial fibrillation, or ventricular pacing. In addition, patients with bundle branch block are excluded from QT variability analyses.The blood draw and processing are standardized. Peripheral blood is collected and allowed to stand at room temperature for 1 hour to minimize the variance in the time from collection to processing. Blood in nonanticoagulated (red‐top) tubes is centrifuged at 1500 rpm for 5 minutes. The serum is then transferred into tubes containing 200‐ to 500‐μL aliquots, frozen in liquid nitrogen and stored at −80°C. Buffy coats are also collected and after a similar low‐speed spin, are transferred into 100‐μL aliquots prior to −80°C storage. Routine clinical laboratory tests are performed and the specific tests determined by the treating physicians. Additional blood is collected for future analysis as part of this study. There are no prespecified blood tests performed on the study samples, but biomarker, metabolomic, and genomic analyses are planned. The specific platforms for genetic testing are not prespecified but will be focused on analyzing large numbers of single‐nucleotide polymorphisms among other things.Device implantation is performed only in patients without clinical evidence of active myocardial ischemia, decompensated heart failure, or infection. All procedures are performed in the standard clinical fashion either with conscious sedation or general anesthesia. In cases of CRT devices, left ventricular pacing leads are targeted to a lateral or posterolateral branch of the great cardiac vein. The final location of the left ventricular pacing lead is assessed using saved fluoroscopic images at the time of device implantation. Toward the end of the procedure, physicians are encouraged to perform programmed stimulation through the device. Programmed ventricular stimulation is performed at 2 drive trains (350 and 500 ms) with up to 3 extrastimuli. The minimum extrastimulus coupling interval is 200 ms. When the ventricular effective refractory period (VERP) is reached for a given extrastimulus, the coupling interval of the preceding extrastimulus is decremented by 10‐ms intervals until VERP is reached or a coupling interval of 200 ms. This process is repeated for all extrastimuli. Morphology and cycle lengths of all sustained ventricular arrhythmias induced are recorded. Data on ICD generator and leads implanted, tachycardia therapy cutoff rates and therapies programmed, use of supraventricular arrhythmia discriminators, and sensitivity settings are also noted.Patients are evaluated every 6 months after implantation and soon after any ICD shock event reported by the patient . At eachunless it was preceded by an ICD shock for supraventricular tachyarrhythmias within a 24‐hour period. We are qualifying this definition because we cannot be absolutely sure that there was no residual effect of the first “inappropriate” shock on the development of the subsequent “appropriate” shock. Deaths are ascertained by phone contact with the next of kin and by searches of the National Death Index. Next of kin interviews regarding the circumstances surrounding the death, death certificates, and medical documentation around the time of death including ICD interrogations whenever possible are collected. This information is subsequently reviewed and adjudicated by 2 independent cardiologists. Each death event is adjudicated on the basis of modified Hinkle–Thaler criteria.13The primary outcome of PROSE‐ICD is ICD shocks for adjudicated ventricular tachyarrhythmias. The secondary outcome is all‐cause mortality. It is expected that some patients will have multiple ICD shocks for ventricular tachyarrhythmias. For the purposes of the analysis, the first confirmed instance will be used Patients are considered part of the cohort after successful ICD implant. For time‐to‐event analysis for the primary end point, patients are censored at the time of death, ICD explantation, left ventricular assist device implantation or heart transplantation, or at the last date of contact among those still alive. For time‐to‐event analysis for all‐cause mortality (secondary end point), follow‐up is censored at the time of ICD explantation, left ventricular assist device implantation or heart transplantation, or at the last date of contact or the date of death in the National Death Index. These events are selected on the basis of their likelihood of altering the patient's underlying cardiac disease process and hence their risk for ICD shocks. The Kaplan–Meier method will be used to estimate the cumulative incidence of these events. Incidence rates for a given event will be calculated by taking the ratio of the total number of events and the total number of person‐years of follow‐up contributed by cohort participants. Hazard ratios of study outcomes adjusted for other participant characteristics will be estimated using proportional hazards regression models.6 the annualized primary end point event rate is estimated at 6%. Annual loss from censoring and losses to follow‐up are estimated at 5%. We calculate that a target enrollment of 1200 patients will result in ≈362 primary end point events by the end of follow‐up and will allow us to detect an estimated hazard ratio of ≥1.85 when comparing extreme quartiles of an independent predictor of ICD shocks with 80% power and a probability of type I error of 5% (2‐sided).On the basis of prior randomized clinical trials,As of December 31, 2011, PROSE‐ICD has enrolled 1177 patients. Although their characteristics are similar to the patients enrolled in MADIT II and SCD‐HeFT, there are some notable differences . OverallAs of December 31, 2011, a total of 234 patients have experienced an ICD shock over an average of 59±23 months of follow‐up. Among them, 143 experienced ICD shocks for ventricular tachyarrhythmias.14 device malfunctions15 and the incidence of inappropriate ICD shocks17 have led some to question whether the benefits outweigh the risks.19 Hence, it is ever more important to accurately identify individuals who stand to benefit the most from this intervention.Sudden cardiac death remains the most common mode of death in the United States and other developed countries. The use of primary prevention ICDs has been shown to reduce total mortality in individuals with severe dilated cardiomyopathy, but only a minority of patients receive appropriate ICD therapies. Furthermore, current rates of periprocedural complications,20 such as microvolt T‐wave alternans,21 magnetic resonance imaging,22 serum‐based biomarkers,23 or a combination of these metrics.24 Additional validation in large cohorts will be required prior to widespread adoption of these techniques in clinical practice.25The optimal strategy for risk stratification of primary prevention individuals at greatest risk for sudden death is still poorly defined. Current strategies rely in large measure on the left ventricular ejection fraction, but limited sensitivity and specificity have led to the proposal of other noninvasive measurementsPROSE‐ICD was conceived with the overall aim of better understanding the biological pathways predisposing to SCD. We will use extensive phenotyping of study participants including genetic, proteomic, metabolomic, electrophysiological, and epidemiological markers to help establish a risk stratification schema that will identify patients at highest and lowest risk for SCD. This study is one of the largest, most extensively phenotyped cohorts of patients undergoing prophylactic ICD implantation. The stringent enrollment criteria and phenotyping accompanied by routine data‐quality checks will provide good internal and external validity of the data.6 there are some differences. First, the percentages of women and nonwhites are 27% and 43%, respectively. This is in contrast to patients from the SCD‐HeFT study, whose enrollment was 23% women and 23% nonwhite subjects. PROSE‐ICD will thus provide insights into the effects of sex and race that cannot be obtained from other studies because of enrollment limitations. Second, the use of heart failure medications at the time of enrollment is optimal, with ≈90% of individuals on β‐blockers and 88% on ACE‐Is. Whether outcomes in patients with ICDs are modulated by optimal heart failure medications remains to be seen. Third, this study reflects current clinical practice patterns and did not require implanting physicians to adopt a prespecified device programming strategy . Hence, the findings from this study stand to better reflect how patients in the general population do over time and provide us with real‐world outcomes. Lastly, the plans for extensive clinical phenotyping and blood collection will allow us to prospectively follow patients and potentially identify novel markers of risk for arrhythmic events.Although the current demographic makeup of this study is similar to previous landmark clinical trials,This study is a prospective, observational study, and inherent limitations of this design as well as variables not currently measured at the time of enrollment may have an effect on the interpretability of the findings. In addition, we have focused on enrollment of individuals with evidence of systolic heart failure. Hence, findings from this study may not apply to the smaller group of individuals with preserved systolic function undergoing primary prevention ICD implantation such as long QT syndrome, early‐stage cardiac sarcoidosis, and other inherited genetic channelopathies. Despite these limitations, we believe that the findings from this study will have significant implications on SCD risk assessment and management as this cohort represents the largest, most extensively phenotyped group of individuals undergoing ICD implantation for primary prevention of sudden death.The PROSE‐ICD study represents a typical cohort of patients receiving ICDs for primary prevention of SCD on the basis of current practice guidelines. Although the clinical demographics appear to be similar to previously published randomized trials, there are some notable differences. PROSE‐ICD will allow us to identify subgroups that stand to benefit the most from these devices and perhaps to craft novel strategies for risk stratification of patients potentially at risk for sudden death.


Obesity is a major public health problem in many poor countries where micronutrient deficiencies are prevalent. A partial meal replacement may be an effective strategy to decrease obesity and increase micronutrient intake in such populations. The objective was to evaluate the efficacy of a partial meal replacement with and without inulin on weight reduction, blood lipids and micronutrients intake in obese Mexican women.2, were allocated into one of the following treatments during 3 months: 1) Two doses/d of a partial meal replacement (PMR), 2) Two doses/d of PMR with inulin (PMR + I) , 3) Two doses/d of 5 g of inulin (INU) and 4) Control group (CON). All groups received a low calorie diet (LCD). Weight, height, hip and waist circumference were measured every 2 weeks and body composition, lipids and glucose concentration and nutrient intake were assessed at baseline and after 3 months.In a randomized controlled clinical trial 144 women (18–50 y) with BMI ≥ 25 kg/mAll groups significantly reduced weight, BMI, waist and hip circumference. Differences between groups were only observed in BMI and weight adjusted changes: At 45 days PMR group lost more weight than INU and CON groups by 0.9 and 1.2Kg, respectively. At 60 days, PMR + I and PMR groups lost more weight than in INU by 0.7 and 1Kg, respectively. Subjects in PMR, PMR + I and INU significantly decreased triglycerides. Energy intake was reduced in all groups. Fiber intake increased in PMR + I and INU groups. Some minerals and vitamins intakes were higher in PMR and PMR + I compared with INU and CON groups.Inclusion of PMR with and without inulin to a LCD had no additional effect on weight reduction than a LCD alone but reduced triglycerides and improved intake of micronutrients during caloric restriction. PMR could be a good alternative for obese populations with micronutrient deficiencies.NCT01505023 There were no significant differences in unadjusted changes between groups. The differences of adjusted changes found were as follows: After 45 days a greater reduction of BMI was observed in PMR group  , and 6%–8% in the long term (> 1 years), when used as part of an overall low-energy diet plan ,12,17,34Treatments with PMR, PMR + I and INU significantly reduced triglycerides after 3 months of treatment. Both groups with inulin and PMR alone had an effect, so this could be attributed to inulin or its combination with a loss of body fat. Of 11 studies reviewed by Delzenne  on the eTotal energy and carbohydrates intakes decreased similarly in all groups and fat intake, including cholesterol, was reduced significantly more in both groups with PMR compared with INU and CON groups. Ditschuneit  found siAn important finding of this study is that PMR could contribute to increase the intake of essential nutrients, which is especially important during caloric restriction and in populations that need to reduce calories but that are at risk of having micronutrient deficiencies. In Mexico for example, there is a high prevalence of obesity, about 30% of adults and 16% of children less than 12 years are obese; and in As it is common in dietary intake data, there was a considerable variability in the nutrient intake data, even when some statistical differences were found, some micronutrient comparisons were low powdered because the sample size was calculated upon body weight. In addition, the study did not find body fat change statistically different between treatments, it has been documented that other methods such as DEXA could have provided more accurate results . Since tIn conclusion, we found no additional effect of including PMR or a PMR added with inulin to a calorie restricted diet on changes in body weight, BMI or fat loss, than a LCD alone; however, intake of PMR, PMR added with inulin or inulin alone contribute to reduce plasma triglycerides during calorie restriction. In addition, treatment of obesity with LCD that includes a PMR significantly increased intake of essential amino acids, vitamins and minerals and contributed to meet daily recommendations. Results of the present study are particular relevant for populations that have increased prevalence of obesity and that at the same time have high prevalence of micronutrient deficiencies. This study points out the importance of using adequately fortified meal replacement products to ensure nutrient adequacy during energy intake restriction for weight loss.PMR: Partial meal replacement; PMR + I: Partial meal replacement with inulin; INU: Inulin; CON: Control group; LCD: Low calorie diet; ANOVA: Analysis of Variance.This study was partially financed by Nucitec SA de CV. MAD was an employee at this company and did not receive any compensation for this study. JLR owns shares of this company. Both participated in the concept and design of the study and providing intellectual interpretation of the results.ART conducted the fieldwork and wrote the initial draft of the paper; MCC contributed with the data analysis and its interpretation as well as with the writing of the manuscript; SG participated in the design of the study and quality assurance; MAD participated in the concept and design of the project; JLR participated in the concept and design and revised critically and intellectually the manuscript. All authors read and approved the final manuscript.