To report the feasibility of intraoperative spectral domain optical coherence tomography (OCT) using the RESCAN 700 for the visualization and evaluation of precut Descemet’s stripping automated endothelial keratoplasty (DSAEK) and prestripped Descemet’s membrane endothelial keratoplasty (DMEK) donor tissue.n = 11), prestripped DMEK (n = 4) preserved in a viewing chamber were examined by intraoperative OCT. Wholly intact donor corneas for penetrating keratoplasty (PK) (n = 8) were also examined as controls. The obtained images were analyzed qualitatively for characteristics of each type of donor.Precut DSAEK (For each type of donor, characteristic images were consistently obtained by intraoperative OCT in both the front and back views through the viewing chamber. In wholly intact donors for PK, appearance of normal corneal curvature and stromal texture with high reflectivity of epithelium and endothelium cell layers were clearly visualized. In precut DSAEK donors, precut lines were characteristically visualized in addition to the intact donor cornea images. In prestripped DMEK donors, identical OCT images to the intact donor cornea were noted when observed from the anterior surface. However, peripheral partial detachments of Descemet’s membrane were characteristically observed in all prestripped DMEK donors when viewed from the back of the viewing chamber.Rapid visualization and rough evaluation of donor tissues for PK, precut DSAEK and prestripped DMEK donor corneas by intraoperative OCT was consistently possible through the viewing chamber. Therefore, this device may be used as an alternative of AS-OCT when the eyebank does not have their own AS-OCT. Although the peripheral detachment in DMEK donors are quite common and clinically non-problematic in DMEK donor quality and subsequent DMEK surgeries, it may be useful to distinguish between wholly intact PK donors and prestripped DMEK donors, enabling to prevent mix-ups of donors, especially when several different types of keratoplasties are scheduled in a same day in one operating theater. Descemet’s stripping automated endothelial keratoplasty (DSAEK) –7 or DesPreviously, anterior segment optical coherence tomography (AS-OCT) has proven useful to screen donor corneas for previous refractive surgeries such as laser in situ keratomileusis , 22. RecHerein, we show the feasibilities of intraoperative spectral domain OCT using the RESCAN 700  , 25 for n = 11) and prestripped DMEK (n = 4) donor corneas from a US eyebank  preserved in a viewing chamber were used. For a control, domestic wholly intact donor corneas taken post-mortem for penetrating keratoplasty (PK) (n = 8) were used. Consent was received for the use of these corneas for this research by the donors’ next of kin. The donor cornea for DSAEK was dissected with a microkeratome  equipped with a 300 μm head at the US eyebank. The DMEK donor graft was also prepared at the US eyebank using the submerged cornea using the backgrounds away (SCUBA) technique [The study was approved by the Ethical Committee of Kanazawa University Graduate School of Medical Science and followed the tenets of the Declaration of Helsinki. Internationally shipped precut DSAEK . The Rescan 700 scan rate was 27,000 axial scans per second. The axial and transverse resolution in tissue are 5 μm and 15 μm, respectively. Each donor tissue was scanned through the transparent window of the corneal viewing chamber, which was placed on a flat table. The OCT images are recorded in a horizontal and vertical orientation. The scan depth was 2 mm and the scan length was adjusted to each donor cornea tissue between 9 and 11 mm (adjustable between 3–16 mm). The surgeon observed the live, uninterrupted intraoperative OCT images during surgery through a small window adjacent to the operating field. Acquired images were evaluated qualitatively for the shape and reflectivity of the donor cornea tissue.For each type of donor , characteristic images were consistently and easily obtained by intraoperative OCT in both the front and back views through the viewing chamber.In wholly intact PK donors, appearance of normal corneal curvature, although not quantitatively measured, and normal stromal texture with high reflectivity of the epithelium and endothelium cell layers were clearly visualized in all donors (n = 8) Fig. .Fig. 1InIn precut DSAEK donors, precut lines were characteristically visualized rapidly in addition to the PK donor cornea images Fig. . Some prIn prestripped DMEK donors, no characteristic images were noted by SD-OCT when observed from the anterior surface, since peripheral image could not be obtained from the anterior surface observation. However, peripheral partial detachments of Descemet’s membrane were characteristically observed in all prestripped DMEK donors when viewed only from the back of the viewing chamber Fig. . Debris All donor corneas were transplanted  to patients, and all corneas remain transparent for at least 12 months.The microscope integrated intraoperative OCT is a relatively new technology. It enables in vivo 3-D visualization of the steps of retina –28 and cHerein, we report the feasibilities of the visualization of donor corneas for PK, precut DSAEK and pre-stripped DMEK by intraoperative OCT using the RESCAN 700. As a result, we showed characteristic images for each type of donor. In wholly intact PK donors, appearance of normal corneal curvature and stromal texture with high reflectivity of the epithelium and endothelium cell layers were clearly visualized. However, the precise corneal curvature could not be measured by RESCAN 700. The images were consistent with those obtained by conventional AS-OCT. In precut DSAEK donors, precut lines were characteristically visualized and the results were also in accordance with those observed by conventional AS-OCT. In prestripped DMEK donors, identical OCT images as the PK donor cornea were noted when observed from the anterior surface. However, peripheral partial detachments of Descemet’s membrane were characteristically observed in all prestripped DMEK donors when viewed from the back of the viewing chamber. Although the peripheral detachment in DMEK donors are quite common and clinically non-problematic in DMEK donor quality and subsequent DMEK surgeries, it may be useful to distinguish between wholly intact PK donors and prestripped DMEK donors by detecting the peripheral detachment sign; it may be difficult or impossible to check the subtle Descemt’s membrane change using slit lamp examination alone.AS-OCT has been used as a routine procedure for tissue evaluation in some advanced eyebanks to detect previous refractive surgeries and corneal scarring . On the One drawback of this study is that quantitative analysis of the donor status is lacking; the quantitative information such as donor thickness or curvature will definitely strengthen this paper. However, RESCAN 700 currently does not have any quantitative capacity. Development of new software in RESCAN 700 that enables quantitative analysis will enhance the capacity of the device.In conclusion, rapid visualization and rough evaluation of donor tissues for PK, precut DSAEK and prestripped DMEK donor corneas by intraoperative OCT was consistently possible through the viewing chamber. Although clinically acceptable, peripheral detachment of Descemet’s membrane of prestripped DMEK donors was successfully and characteristically observed by intraoperative OCT. It may be possible to prevent mix-ups of donors by checking those donors just before surgery by surgeon especially several different types of keratoplasties are scheduled in a same day in one operating theater.


During ageing, total testosterone (TT) declines and SHBG increases, resulting in a greater decline of free testosterone (FT) compared to TT. However, guidelines suggest using TT to diagnose androgen deficiency and to reserve FT only for men with borderline TT. We investigated if isolated low FT or isolated low TT was associated with androgen-related endpoints in healthy men.3369 community-dwelling men, aged 40-79, were included. We assessed differences between men with both normal TT (=10.5 nmol/L) and calculated FT (=220 pmol/L) (referent), men with normal TT/low FT (group 1) and men with low TT/normal FT (group 2) by descriptive statistics and ordinal logistic regression adjusted for age, centre, BMI and comorbidities.2540 men had normal TT (18.4-5.5 [mean-SD] nmol/L) and FT (326-75 pmol/L). There were 261 men in group 1 , low FT (195-22 pmol/L)) and 92 men in group 2 (low TT (9.6-0.7 nmol/L), normal FT (247-20 pmol/L)).Compared to referent, men in group 1 were older and had higher SHBG, whereas group 2 was younger and had lower SHBG. Men in group 1, but not group 2, were in poorer health, had lower haemoglobin and a decrease in bone ultrasound measurements. Regression analysis showed that men in group 1 had less frequent morning erections, more erectile dysfunction, fewer sexual thoughts and more physical symptoms . Compared to referent, sexual and physical symptoms did not differ in group 2.Independent of age, BMI and comorbidities, men with isolated low FT, but normal TT, have more androgen deficiency-related symptoms than men with normal TT and FT levels; whereas symptoms do not differ in men with isolated low TT. Not only total, but also FT levels should therefore be assessed in men with hypogonadal symptoms.


Matthew Allen and Peter Reese argue that evidence-based efforts should be implemented to expand living kidney donation. Despite efforts to expand access to organ transplantation worldwide, in many countries a majority of patients face several years of waiting for a kidney to become available.While governments and the transplant community actively encourage people to become organ donors after death, analogous campaigns expanding awareness of living donation are almost nonexistent.Research suggests altruism depends on a social context that actively encourages giving behavior.Transplant centers should professionalize the process of helping patients find a donor and employ evidence-based approaches to increasing living organ donation.The community should launch a campaign disseminating information about the risks and benefits of living kidney donation and encouraging the public to consider donating a kidney; this campaign should have the same energy and creativity that characterizes the efforts supporting deceased organ donation.If you needed a kidney transplant, would anyone donate to you? For most people, the answer is “no.” Millions of individuals are eligible to donate a kidney, but relatively few become donors. In the United States, there are 100,000 people on the waiting list for a kidney transplant who spend years hoping for an offer. Meanwhile, 6% die annually, patients and families suffer the daily hardships of dialysis, and Medicare spends billions of dollars per year on their care [Deceased donor transplantation (DDT) is a well-funded, organized enterprise. In many countries, DDT relies on a network of transplant centers and nonprofit organizations to coordinate organ donation, transportation, and transplantation. Trained professionals approach next-of-kin to lead delicate discussions about organ donation. National campaigns promote donor registration using billboards, public events, and targeted advertising strategies.In comparison, initiatives promoting LKD are limited in scope. This is problematic given that 1) efforts to optimize DDT have not kept pace with demand and face substantial barriers ; 2) liviMoreover, patients who seek a donor usually have little support or guidance. If addressed at all, talking about LKD commonly involves a nephrologist asking whether the patient knows a donor. Most physicians lack training in advising patients about approaching potential donors, and this task is difficult in every sense. Whom should you ask? What should you say? The uncertainty can be overwhelming. Many patients understandably avoid asking, instead choosing to wait for a kidney from a deceased donor.Why are deceased and living donor transplantation managed so differently? One problem is how we have conceptualized LKD. The decision to donate is regarded as a private, mysterious act. When considering a “nondirected” donor—someone who wants to donate with no particular recipient in mind—a skeptical transplant community has historically expressed concerns that many such individuals were mentally ill . In the While the US government maintains a website promoting organ donation, it focuses almost exclusively on DDT. The site features public service announcements for television, radio, and print encouraging donor registration. It includes “tool kits” to facilitate promotional campaigns at hospitals, campuses, and offices. Analogous materials for LKD are notably absent, reflecting policy not to promote living donation because the long-term risks are not fully known . ImportaIn addition, ethical concerns within the transplant center have fostered a “hands off” approach to expanding LKD . PhysiciAlthough financial incentives have long been the focus of discussions about increasing LKD, ethical objections make widespread adoption of payment for donation unlikely. Concerns regarding unjust inducement may be specially pronounced in developing countries, where economic vulnerability is widespread. We suggest an approach that avoids the associated legal and ethical concerns. Research on altruism suggests it requires not only individual motivation but also a social context that actively encourages generosity. For example, in diverse circumstances, the most consistent determinant of giving behavior is a person being asked . What stWe propose reconceptualizing LKD based on knowledge of donor motivations, barriers to donation, and the nature of altruistic behavior. We envision a strategy that includes 1) initiatives from within transplant centers to professionalize the process of helping patients find a donor and 2) a campaign from outside the transplant enterprise disseminating information about donation’s risks and benefits and encouraging the public to consider donating a kidney .IReconceptualizing LKDRecognizing ESRD as a shared public health problemDemystifying the act of donation by listening to donor motivations and experiencesReframing donation as an important form of service to one’s communityAcknowledging that risk to donors does not preclude rational decision making or informed consentUnderstanding that altruistic behavior depends on social contexts that actively encourage itIIEngaging the Public on LKDHarnessing creativity and expertise in media to engage the public on LKDEducating the public about the risks and benefits of donationDisseminating compelling stories about kidney recipients and their donorsCountering portrayals in the media suggesting donation is exoticEncouraging the public to consider donating a kidney, with approaches addressing both directed donation  and nondirected donation to a strangerLiving kidney donorsLiving kidney transplant recipientsPatient advocacy groupsEliciting the input and involvement of diverse stakeholders in the management of the proposed campaign:IIIOvercoming Barriers to LKDEducating transplant candidates about the risks and benefits of LKD transplantationCoaching patients about how to raise the subject of donation with potential donorsFinding a “champion” to approach potential donors on the patient’s behalf (when appropriate and feasible)Training professionals to empower patients in their search for a donor:Developing personalized and targeted strategies to address culture and race-specific barriers to LKD transplantMinimizing financial barriers to donation by reimbursing expenses related to donation, including the process of donor evaluation and lost wagesIVExpanding Protections for Donors and Promoting Ethical StandardsTransparency about risks of donationAssessments of donor understandingUse of independent donor advocatesPsychosocial evaluationMaintaining commitment to robust informed consent practices at the center level:Establishing legal protections for living donors that ensure eligibility for insurance coverage without increased premiumsEstablishing regulations preventing donors from bearing costs of treatment for complications of donationTransplant tourismOrgan traffickingCommercializationDonation from minorsReaffirming commitment to combating unacceptable transplant practices:Addressing root causes of organ trafficking by improving access to legal and ethically appropriate organ transplantationTransplant centers should develop an infrastructure for employing evidence-based interventions to optimize LKD . TrainedWhile transplant professionals play a central role in expanding LKD, a transformative approach entails efforts from beyond the transplant center. Foundations and nonprofits should launch a campaign to educate the public about the risks and benefits of LKD. Enlisting expertise in media could spread awareness of the dire need for transplants and engage people in ways that prose and statistics cannot. As opposed to an abstract, far-fetched possibility, organ donation should be something we are reminded of daily—a commendable act we should all be encouraged to consider. Given many governments’ risk-averse stance on LKD and the need for transplant centers to avoid conflicts of interest, foundations and nonprofit organizations must lead this effort.Reasonable objections include the fact that kidney donation has risks, including perioperative death , surgical complications, and an increased risk of developing ESRD  . Donor rMost of us are fortunate and will never need to hope for a donor to come forward. Given the expanding need for transplants and the human toll of kidney disease, it is incumbent on us to explore new approaches to expanding kidney donation. By promoting informed decision-making, celebrating donors, and removing barriers to donation, perhaps we can collectively bring kidney transplantation to all who need it.


The optimal penetration of antiretroviral agents into central nervous system (CNS) may be a balance between providing adequate drug exposure to inhibit HIV-replication whilst avoiding concentrations associated with toxicities.Cerebrospinal-fluid (CSF) exposure of efavirenz and metabolites 7-hydroxy (7OH-) and 8OH-efavirenz were assessed after at least 12 weeks of antiretroviral therapy in HIV-infected subjects randomized to commence antiretroviral regimens containing efavirenz at either 400 mg or 600 mg once daily. Clinical, pharmacokinetic and pharmacogenomic factors associated with CSF efavirenz and its metabolite concentrations were assessed.reference) in 11/14 and 7/14 subjects randomized to the 400 mg and 600 mg doses of efavirenz, respectively. Whilst CSF efavirenz concentration was significantly associated with plasma concentration (P<0.001) and CYP2B6 genotype , CSF 8OH-efavirenz concentration was not . Lastly, CSF 8OH-efavirenz concentration was associated with efavirenz symptom questionnaire results at one year .Of 28 subjects who completed all study procedures (14/14 on efavirenz 400 mg/600 mg), CSF HIV RNA was below 20 copies/mL in all at the time of examination. Concentrations of efavirenz and 7OH-efavirenz in the CSF were slightly lower when dosed at 400 mg versus 600 mg, although this was not statistically significant. A different trend was observed regarding 8OH-efavirenz concentrations where CSF exposure was slightly increased in the 400 mg efavirenz arm see . EfavireWith both doses of efavirenz studied, CSF concentrations were considered adequate to inhibit HIV-replication, although concentrations of 8OH-efavirenz were greater than that reportedly associated with neuronal toxicity. CSF exposure of 8OH-efavirenz was not dependent on plasma exposure and we postulate may be subject to saturable pharmacokinetic effects.


Transposable elements (TEs) allow rewiring of regulatory networks, and the recent amplification of the ISX element dispersed 77 functional but suboptimal binding sites for the dosage compensation complex to a newly formed X chromosome in Drosophila. Here we identify two linked refining mutations within ISX that interact epistatically to increase binding affinity to the dosage compensation complex. Selection has increased the frequency of this derived haplotype in the population, which is fixed at 30% of ISX insertions and polymorphic among another 41%. Sharing of this haplotype indicates that high levels of gene conversion among ISX elements allow them to ‘crowd-source’ refining mutations, and a refining mutation that occurs at any single ISX element can spread in two dimensions: horizontally across insertion sites by non-allelic gene conversion, and vertically through the population by natural selection. These results describe a novel route by which fully functional regulatory elements can arise rapidly from TEs and implicate non-allelic gene conversion as having an important role in accelerating the evolutionary fine-tuning of regulatory networks.DOI:http://dx.doi.org/10.7554/eLife.05899.001 Mutations change genes and provide the raw material for evolution. Genes are sections of DNA that contain the instructions for making proteins or other molecules, and so determine the physical characteristics of each organism. Genetic mutations that increase an organism's number of offspring and chances of survival are more likely to be passed on to future generations. Changes to when or where a gene is switched on  can also provide fitness benefits and can therefore be selected for during evolution.Transposable elements are sequences of DNA that are also called ‘jumping genes’ because they can make copies of themselves and these copies of the transposable element can move to other locations in the genome. Some transposable elements contain sequences that switch on nearby genes. If different copies of a transposable element that contains such a regulatory sequence insert themselves in more than one place, it can result in a network of genes that can all be controlled in the same way. The regulatory sequences contained within transposable elements are not always optimal, but they can be fine-tuned through evolution.Drosophila miranda has a transposable element called ISX that has, over time, placed up to 77 regulatory sequences around one of this species' sex chromosomes. Just as in humans, female flies are XX and males are XY; but having only one copy of the X chromosome means that male flies need to increase the expression of certain genes to produce a full-dose of the molecules made by the genes. This process is called dosage compensation and in 2013 the 77 ISX regulatory sequences on the fruit fly's X chromosome were shown to help recruit the molecular machinery that carries out dosage compensation to nearby genes, albeit inefficiently. Now Ellison and Bachtrog—who also conducted the 2013 study—report how these transposable elements have been fine-tuned to make them more effective for dosage compensation.A fruit fly called Ellison and Bachtrog uncovered two mutations that make the ISX transposable element better at recruiting the dosage compensation molecular machinery. ISX spread around different locations along the fly's X chromosome before these mutations arose; this means that initially none of the 77 insertions carried the two mutations, but now 30% of the 77 elements have the mutations in all flies, and 41% have them in only some flies.The same mutations have spread between the different ISX elements because transposable elements with the mutations have been used to directly convert other ISX elements without them. These mutations have also become more common in the fruit fly population by being passed on to offspring and increasing their survival. These two routes have accelerated the fine-tuning of these transposable elements for use in gene regulation. This implies that regulatory sequences derived from transposable elements evolve in a way that is fundamentally different from those that arise by other means, as the direct conversion between these insertions allows fine-tuning mutations to spread more rapidly.DOI:http://dx.doi.org/10.7554/eLife.05899.002 A substantial portion of animal genomes is composed of repetitive sequences, including gene duplicates, satellite DNA, and transposable elements. Gene conversion is a major force shaping the evolution of repetitive regions, and interlocus or non-allelic gene conversion between sequence duplicates has been studied extensively for its role in concerted evolution . Non-allDrosophila miranda was driven by TE-mediated amplification of a functional but suboptimal binding motif  that binds to a GA-rich sequence motif (the MSL recognition motif) at a number of chromatin entry sites on the X chromosome . We prevrent age . XL is hon years ; chromoson years , and theon years . Dosage on years . Intereson years .D. miranda and is derived from the abundant ISY element. Compared to ISY, ISX contains a 10 basepair deletion that creates a MSL recognition motif, thereby allowing it to act as a chromatin entry site . The haplotype consists of two mutations (G → T and A → T), separated by two basepairs, which are in perfect linkage disequilibrium among ISX but not ISY elements . BecauseD. miranda strain MSH22  from in MSH22 . We compin MSH22 .10.7554/D. melanogaster genome results in recruitment of the MSL complex to an ectopic autosomal location (D. melanogaster chromosome 2L at cytosite 38F1 using recombinase mediated cassette exchange (RMCE)  . We thenInterestingly, each T allele, when assayed separately, has a negative effect on MSL binding levels compared to the ancestral G or A allele . HoweverIt is unlikely that the TT haplotype arose multiple times by parallel mutation, and there are two possibilities that could explain its prevalence among MSH22 ISX insertions. First, this double mutation may have occurred early during the process of ISX amplification, thus giving rise to two lineages of ISX: one that carries the ancestral GA haplotype, and the other with the TT haplotype. The TT-harboring elements in MSH22 would then all be descendants from the latter ISX lineage. Alternatively, this mutation may have occurred only after the GA-containing ISX element was fixed in the population at all 77 neo-X insertion sites, at which point it was spread among independent ISX elements via non-allelic gene conversion.D. miranda. A canonical signature of non-allelic gene conversion is the presence of shared polymorphisms across sequence duplicates . While many of these haplotypes are likely to be neutral or deleterious, some may be beneficial, as in the case of the TT haplotype. Such a scenario avoids the waiting time for a double mutation, as well as the fitness valley that would have to be traversed if the two mutations were to occur sequentially on the ISX background.Our transgenic experiments show that each individual T allele actually decreases the binding affinity for the MSL complex relative to the ancestral GA haplotype. Thus, TA or GT haplotypes should be selected against in the population if present on a functional ISX element. Consistent with the deleterious effect of individual T alleles, the TA and GT haplotypes are present on some ISY elements but completely absent from ISX, that is, we find the two T mutations to be in perfect linkage disequilibrium among ISX elements but not ISY . While mTo conclude, our findings suggest that TE-dispersed binding motifs follow an evolutionary trajectory that is fundamentally different from those that arise by other means. The complementary roles of TEs in dispersing regulatory motifs, and gene conversion in spreading subsequent refining mutations, combine to allow for the rapid rewiring and fine-tuning of gene regulatory networks. This process adds a new layer of complexity onto how TEs influence regulatory innovation, as well as a new context in which gene conversion affects genome evolution.Isofemale lines were established from individuals collected in Northern California and inbred for several generations. DNA was extracted from 1–8 females per line using the Qiagen PureGene kit (Netherlands) and fragmented by nebulization. Paired-end Illumina libraries were constructed using standard protocols  and sequD. miranda MSH22 reference assembly (GenBank: AJMI00000000.2) using bowtie2 (https://github.com/chris-ellison/MSAvariants) was used to identify sequence variants within the alignments. We also PCR amplified eight of the ISX insertions where the TT haplotype was polymorphic. We confirmed that this polymorphism was present at each of these insertions and estimated the base-calling accuracy of the assemblies by sequencing the PCR products using Sanger technology.Resequencing data were mapped to version 2.2 of the  bowtie2 . ISX loc bowtie2 . Contigs bowtie2  and visu bowtie2 . A custoD. melanogaster embryos carrying a RMCE landing site at cytosite 38F1 on chromosome 2L (Bloomington Drosophila Stock Center strain #27388). Transformants were verified by PCR and Sanger sequencing.We used the QuikChange Lightning site-directed mutagenesis kit from Agilent Technologies  and the ISX element cloned in Male third instar larvae (∼250 mg) were collected from F1 hybrids between ISX-GA and each of the other three engineered lines: ISX-GT, ISX-TA, and ISX-TT. Chromatin immunoprecipitation was performed for four biological replicates of each of these lines using the MSL2 d-300 primary antibody from Santa Cruz Biotechnology Inc.  and the protocol described in To determine if the TT frequency among neighboring ISX elements was correlated, we clustered elements within 100 kb of each other and calculated the standard deviation in TT allele frequency within clusters. We then compared these values to 1000 permutations where TT allele frequency was randomly shuffled between ISX locations.The haplotype diversity and allele frequency spectrum resampling tests were performed by drawing, without replacement, two groups of size 617 and 674, respectively, from the pool of 1291 ISX sequences. The intergroup difference in haplotype diversity, as well as the number of derived variants with frequency of 0.75 or greater, was calculated for each of 1000 replicates and compared to the difference between the TT and GA groups. review process). Similarly, the author response typically shows only responses to the major concerns raised by the reviewers.eLife posts the editorial decision letter and author response on a selection of the published articles . An edited version of the letter sent to the authors after peer review is shown, indicating the substantive concerns or comments; minor concerns are not usually shown. Reviewers have the opportunity to discuss the decision before the letter is sent  and 3 reviewers, one of whom is a member of our Board of Reviewing Editors. One reviewer, Hideki Innan, has agreed to reveal his identity.Thank you for sending your work entitled “Transposable elements crowd-source mutations to rapidly fine-tune regulatory networks” for consideration at The Reviewing editor and the other reviewers discussed their comments before we reached this decision, and the Reviewing editor has assembled the following comments to help you prepare a revised submission.There was general agreement that this is an exciting paper, and a substantial contribution to how our understanding of how regulatory evolution might occur. We also thought you could relatively easily do more to strengthen the case for selection having acted to increase the frequency of the TT haplotype. Simple tests based on the extent of haplotype sharing, the allele frequency distribution, etc., appear not to have been carried out. There is an obvious a priori hypothesis to test here, and multiple loci to test it with.There was also general agreement that your title, albeit catchy, is not professional enough. Perhaps use this phrase in the discussion instead, and use a more descriptive title here. How about something like “Non-allelic gene conversion between repetitive sequences enable rapid evolutionary change at multiple regulatory sites encoded by transposable elements”?Reviewer #2 minor comments:D. miranda strain MSH22”, it seems that the TT haplotype already existed in the ISY family, and so did the GT and TA haplotypes? But because they were rare, the authors assumed that the TT haplotype is derived? The very origin of ISX was GA, and then TT was transferred from ISX to ISY? Do I understand correctly? More detailed description would be nice.1) It is a bit unclear on the origin of the TT holpotype. According to the description in the beginning of the subsection “MRE variation among ISX insertions in 2) I found an interesting (but not very emphasized) observation that the two sites are in strong LD in ISX, but not in ISY. A small sub-table or something in 3) I like that the authors looked at polymorphism. What does the frequency spectrum of TT  I wonder how the spatial distribution of shared sites across the TE region looks. Does your hypothesis predict an excess of shared sites around the two sites? Maybe so, but I'm not sure. Once the TT haplotypes was introduced in the ISX family, TT haplotypes may be preferentially transferred within the neo-X. In such a case, gene conversion does not matter… I don't know any theory that handles such a complicated case.Reviewer #3 minor comments:In the beginning of the subsection “Non-allelic gene conversion is spreading the TT haplotype among ISX insertions”: Actually the decreased linked variation in the TT haplotype is more likely to be the result of non-allelic gene conversion, rather than selection. Gene conversion may happen across hundreds nucleotides, and this in turn can be responsible of the observed decreased variation, on the scale of course of a hundreds bp around the TT sites. There was general agreement that this is an exciting paper, and a substantial contribution to how our understanding of how regulatory evolution might occur. We also thought you could relatively easily do more to strengthen the case for selection having acted to increase the frequency of the TT haplotype. Simple tests based on the extent of haplotype sharing, the allele frequency distribution, etc., appear not to have been carried out. There is an obvious a priori hypothesis to test here, and multiple loci to test it with.There was also general agreement that your title, albeit catchy, is not professional enough. Perhaps use this phrase in the discussion instead (to encourage popular science writers to pick it up…), and use a more descriptive title here. How about something like “Non-allelic gene conversion between repetitive sequences enable rapid evolutionary change at multiple regulatory sites encoded by transposable elements”?We have strengthened the case for selection having acted to increase the frequency of the TT haplotype, and in addition to haplotype diversity include two additional tests of directional selection using the allele frequency spectrum and nucleotide diversity. Consistent with models of directional selection, the frequency spectrum of TT-carrying haplotypes shows an excess of high frequency derived alleles, compared to the GA-carrying ISX elements, and nucleotide diversity is reduced at ISX elements where the TT haplotype is fixed, relative to GA-containing insertion sites. The results of the three statistical tests of selection are now presented in the new Reviewer #2 minor comments:1) It is a bit unclear on the origin of the TT holpotype. According to the description in the beginning of the subsection “MRE variation among ISX insertions in D. miranda strain MSH22”, it seems that the TT haplotype already existed in the ISY family, and so did the GT and TA haplotypes? But because they were rare, the authors assumed that the TT haplotype is derived? The very origin of ISX was GA, and then TT was transferred from ISX to ISY? Do I understand correctly? More detailed description would be nice.There are multiple possibilities of how the TT haplotype could have originated—either by a double mutation in a GA-containing ISX element, or the two mutations might have occurred subsequently , or it might have originated in an ISY element, and gene converted onto ISX. We cannot distinguish between these possibilities, but add some discussion on its origin in the Discussion section of the manuscript.2) I found an interesting (but not very emphasized) observation that the two sites are in strong LD in ISX, but not in ISY. A small sub-table or something inmight help to convince readers.We added a second panel to 3) I like that the authors looked at polymorphism. What does the frequency spectrum of TT  look like? Is it what expected under a directional selection model? Are there any sites that are not fixed in the population yet, but there is already GA/TT polymorphism?We have added a supplementary figure  that compares the frequency spectrum across TT-carrying ISX sequences versus GA-carrying ISX sequences. Consistent with directional selection models of incomplete hitchhiking due to recombination and/or gene conversion, the frequency spectrum of TT-carrying haplotypes shows an excess of high frequency derived alleles, compared to the GA-carrying ISX elements.4) I wonder how the spatial distribution of shared sites across the TE region looks. Does your hypothesis predict an excess of shared sites around the two sites? Maybe so, but I'm not sure. Once the TT haplotypes was introduced in the ISX family, TT haplotypes may be preferentially transferred within the neo-X. In such a case, gene conversion does not matter… I don't know any theory that handles such a complicated case.The spatial distribution of shared polymorphism across the ISX element is shown on the bottom right of Reviewer #3 minor comments:In the beginning of the subsection “Non-allelic gene conversion is spreading the TT haplotype among ISX insertions”: Actually the decreased linked variation in the TT haplotype is more likely to be the result of non-allelic gene conversion, rather than selection. Gene conversion may happen across hundreds nucleotides, and this in turn can be responsible of the observed decreased variation, on the scale of course of a hundreds bp around the TT sites.We don't think this is the case. Non-allelic gene conversion can certainly reduce variation among insertions. However, while non-allelic gene conversion may decrease variation within the TT haplotype, it should equally decrease variation within the GA haplotype (assuming that they are subject to similar levels of gene conversion). The test of selection we employed shows that given its frequency, the TT haplotype harbors significantly less variation than the GA haplotype . We now also show that the frequency spectrum of TT-carrying haplotypes shows an excess of high frequency derived alleles, compared to the GA-carrying ISX elements , which further strengthens the case that selection is operating on the TT haplotype.: The top-to-bottom orientation of the sequence in this figure to me is kind of counter intuitive, and is different from the more usual left-to-right orientation in. I thinkmight work better if rotated 90° counterclockwise; the authors can try and see if they also like it better.


Some grafts harvested from the groin area do not become hyperpigmented and in an individual with multiple pieces of grafts, the hyperpigmentation of the pieces may vary. This study evaluates the orientation of the inset of groin grafts according to their donor sites (superior-inferior) and its role in graft hyperpigmentation.Patients with hand burn contracture or syndactyly who required at least 2 pieces of grafts were enrolled. In each patient, one piece of the graft was inset in the same direction of the orientation of the donor site and the other in the opposite direction. Six months after the operation, the pigmentation was scored by a subjective scaling from 1 to 5. Thirty-four fingers of 15 patients were included. The mean grade of pigmentation in grafts inset in the direction of the donor site was 3.00±1.118 and in those inset in the opposite direction was 2.88±1.409. This was not statistically significant.Our findings revealed that although the grafts inset in the opposite direction of the donor site, they were less pigmented. So orientation of full thickness groin grafts did not affect hyperpigmentation of the grafts in burn contracture and syndactyly hands. STSG are more vulnerable to contractures than FTSG2-5 and are more likely to require revisions.6-8 Although the results of studies are not consistent.9 The donor site of the STSG may be more annoying to the patient than the FTSG that is closed primarily. On the other hand, FTSGs may become hyperpigmented and hair growth may be an annoying problem with them.10-13 Theoretically, from the view of both long term function and appearance, the best option would be a FTSG without hyperpigmentation and hair growth.Release of hand burn contractures generally leads to deficiency of skin that necessitates skin grafting. Release of syndactyly hands, especially when the webbing is beyond the proximal interphalangeal joint, usually requires skin grafting.The most popular donor site for the harvest of the FTSG is the groin area. The groin area is a good source for large grafts, it is easily closed primarily, and its scar is inconspicuous. Full thickness graft harvest from the groin is almost easy. On the other hand, FTSG harvested from the groin is vulnerable to hyperpigmentation and hair growth. In practice, some FTSGs harvested from the groin area do not become hyperpigmented. Even in an individual patient who needs multiple pieces of graft, some pieces of FTSGs do hyperpigment and others do not. FTSGs used in the hand are usually in small pieces. During their inset, the orientation of the grafted skin according to its original site (superior-inferior) is not mentioned. This may be a potential cause for varied hyperpigmentation. This study is planned to examine whether the orientation of the inset of FTSGs harvested from the invaluable donor site of the groin, according to their donor sites, has a role in the graft hyperpigmentation. P value of <0.05.Patients with burn contracture of fingers or syndactyly who required at least 2 pieces of FTSGs were included in the study. All the grafts were harvested elliptically from the groin area by the first, second and the last authors with a No.15 scalpel while the assistant exerted counter traction over the skin. After harvest, the graft was defatted and special attention was made to remove all hair follicles from the dermis of the graft. Each patient was the control of herself/himself. In every patient, one piece of the graft was inset in the same direction of the orientation of the donor site and the other piece in the opposite direction. The orientation of the graft in each finger was randomly determined before the operation and was not affected by the conditions of the operation. The grafts were inset with the similar tensions. Grafts were covered with tie-over dressings that were remained in their place for 5 days. Among the patients operated in the period between December 2009 to June 2011, 34 fingers  in 15 patients were studied. Grafts were checked according to the pigmentation by the third author 6 months after the operation while she was blind about the orientation of the grafts. The pigmentation of each piece of the graft was scored by a subjective scaling from 1 to 5 in which 1 was very mild pigmentation and 5 were very severe pigmentation cases. Comparison of the mean grade of hyperpigmentation was done with Mann-Whitney U test and the statistical significance was set at a Fifteen patients were enrolled in the study. In these patients, 34 fingers were studied. The average age of the 15 patients was 17.4 years . Eight patients were female and 7 were male. The pathologic process necessitating grafting in the fingers was burn in 11 patients and syndactyly in 4 patients . In one patient, the index and the middle fingers of one hand were compared with each other and the ring and little fingers of the same hand were compared separately. In another patient, 2 fingers of one hand were compared with each other and 2 fingers of the other hand were compared separately. In conclusion, we had 34 cases: 17 grafts that were inset in the direction of the donor site and were compared with 17 grafts that were inset in the opposite direction of the donor site. In the total 34 cases, the pathologic process was burn in 26 cases (76.5%) and syndactyly in 8 cases (23.5%). P=0.734).The mean grade of the pigmentation of the grafts that were inset in the direction of the donor site was 3.00±1.118 and the mean grade in the grafts that were inset in the opposite direction of the donor site was 2.88±1.409. The mean grade of hyperpigmentation was not significantly different in the two groups (Full-thickness skin grafting is a valuable treatment for release of finger burn contractures and syndactylies. On the other hand, hyperpigmentation of full-thickness grafts is an annoying problem to the patient. It does not occur uniformly in all patients and we cannot predict its occurrence and its severity before the operation. 14 plantar instep,15-23 and so on.24-26 All of these alternative sites have some disadvantages; the most common of them are poor scars and shortage of the available skin. Treatment of hyperpigmented palmar grafts is discussed in some studies.16,26,27 But, the prevention is better than the treatment.The only factor that is known to influence hyperpigmentation of the skin grafts is the site of the donor area. This has led to search for other donor sites such as upper arm, ulnar aspect of the wrist,28-30 But, to our knowledge, there is no study about the effect of the orientation of the skin grafts. Every effort should be done to find the factors related to the occurrence of hyperpigmentation and to decrease its chance or its severity. Orientation of some types of grafts is very important, e.g., vein and nerve grafts.In this study, it was shown that although the grafts inset in the opposite direction of the donor site were less pigmented, the difference was not statistically significant. But, it opens a search for identifying factors that may have a role in the hyperpigmentation of the grafts and probably predicting or decreasing its occurrence or severity. In conclusion, the orientation of full thickness groin grafts does not affect hyperpigmentation of the grafts in burn contracture and syndactyly hands.The authors declare no conflict of interest.


APOE-ɛ4. These findings reveal that elevated brain iron adversely impacts on AD progression, and introduce brain iron elevation as a possible mechanism for APOE-ɛ4 being the major genetic risk factor for AD.Brain iron elevation is implicated in Alzheimer's disease (AD) pathogenesis, but the impact of iron on disease outcomes has not been previously explored in a longitudinal study. Ferritin is the major iron storage protein of the body; by using cerebrospinal fluid (CSF) levels of ferritin as an index, we explored whether brain iron status impacts longitudinal outcomes in the Alzheimer's Disease Neuroimaging Initiative (ADNI) cohort. We show that baseline CSF ferritin levels were negatively associated with cognitive performance over 7 years in 91 cognitively normal, 144 mild cognitive impairment (MCI) and 67 AD subjects, and predicted MCI conversion to AD. Ferritin was strongly associated with CSF apolipoprotein E levels and was elevated by the Alzheimer's risk allele,  Brain-iron elevation is implicated in Alzheimer's disease (AD), but the impact of the metal on disease outcomes has not been analysed in a longitudinal study. Here, the authors examine the association between the levels of ferritin, an iron storage protein, in the cerebrospinal fluid (CSF) of AD patients and show that CSF ferritin levels predict AD outcomes. Cortical iron elevation is increasingly reported as a feature of Alzheimer's disease (AD)CSF ferritin could be an index of brain iron load. Ferritin is the iron storage protein of the body and is elevated in AD brain tissue5679141–42 ratio and apolipoprotein E (ApoE) levels, in predicting various outcomes of AD. However, the nature of the relationship between CSF ferritin levels and cognitive performance was different from the other biomarkers, and, in contrast, CSF ferritin appears as a trait variable, and not a marker of disease.Here, we examined the association of baseline CSF-ferritin data with biomarker, cognitive, anatomical and diagnostic outcomes over 7 years in the Alzheimer's disease Neuroimaging Initiative (ADNI) prospective clinical cohort. We show that CSF ferritin levels have similar utility compared with more established AD CSF biomarkers, the tau/Aβn=91), mild cognitive impairment  and AD (n=67) subjects , CSF ferritin levels were predicted by Aβ1–42  and tau  alleles are the major genetic risk for AD1-42 , which was improved when Aβ1–42 and tau (non-significant terms) were removed from the model , with the major AD risk allele, ɛ4, inducing 22% higher levels than non-ɛ4 carriers  compared with non-ɛ4 carriers  and tau/Aβ1–42 ratio (P=0.025), independently, in a multiple regression model containing the AD biomarkers and other clinical variables. In tertile analysis, high (>7.2 ng ml−1), compared with low (<5.4 ng ml−1), levels of ferritin were associated with an ≈3 point poorer ADAS-cog13 score  score was associated with CSF ferritin P=0.006; , ApoE le13 score . Similar13 score  were assAS-Cog13 , as prevP=0.006) and tau/Aβ1-42 ratio (P=2.70 × 10−7) were still associated with rate of cognitive change (interacted with time), as previously reported21P=4.93 × 10−4 main effect only; To determine whether baseline values of CSF ferritin predict longitudinal cognitive outcome, we constructed a mixed effects model of annual ADAS-Cog13 scores over 7 years , but there was only a trend for ApoE (P=0.066). Hence, in both cognitive scales, CSF ferritin impacted on performance by a constant amount, regardless of disease status, thus appearing as a trait variable and not a marker of disease.We additionally modelled cognition using the Rey verbal learning test (RAVLT), which is more sensitive in distinguishing control and MCI patients1-42 measurements. In a minimal model  against the values for the quintile boundaries of each analyte . The gradient of these functions estimates the change in mean age of conversion (in months) associated with one unit change in the baseline CSF analyte. For comparison between biomarkers, we also expressed the change in mean age of conversion associated with a s.d. change to the analyte value. One s.d. change to ferritin was associated with a 9.5-month shift in age of conversion, compared with 18.2 and 8.6 months for ApoE and tau/Aβ1–42, respectively , and decreased the odds for ApoE . Including all three analytes into the one model increased the predictive value of each analyte (OR (CI): ferritin=2.32  and tau/AβAPOE ɛ4 genotype) was 0.6079  of the base model  has an epistatic interaction with APOE ɛ4 to increase AD risk and accelerates disease onset by 5.5 years  database (adni.loni.usc.edu). The ADNI study has been previously described in detailwww.adni-info.org.The principal investigator of this initiative is Michael W. Weiner, MD, VA Medical Center and University of California, San Francisco. ADNI is the result of efforts of many co-investigators from a broad range of academic institutions and private corporations, and subjects have been recruited from over 50 sites across the United States and Canada. The initial goal of ADNI was to recruit 800 subjects but ADNI has been followed by ADNI-GO and ADNI-2. To date these three protocols have recruited over 1,500 adults, ages 55 to 90, to participate in the research, consisting of cognitively normal older individuals, people with early or late MCI, and people with early AD. The follow-up duration of each group is specified in the protocols for ADNI-1, ADNI-2 and ADNI-GO. Subjects originally recruited for ADNI-1 and ADNI-GO had the option to be followed in ADNI-2. For up-to-date information, see Inclusion criteria were as follows: (1) Hachinski Ischaemic Score ≤4; (2) permitted medications stable for 4 weeks before screening; (3) Geriatric Depression Scale score<6; (4) visual and auditory acuity adequate for neuropsychological testing; good general health with no diseases precluding enrolment; (5) six grades of education or work history equivalent; (6) ability to speak English or Spanish fluently; (7) a study partner with 10 h per week of contact either in person or on the telephone who could accompany the participant to the clinic visits.Criteria for the different diagnostic groups are summarized in 1–42 and tau levels in CSF were measured using the Luminex platform. ApoE and ferritin protein levels were determined using a Myriad Rules Based Medicine platform . CSF Factor H (FH) levels were measured using a multiplex human neurodegenerative kit  according to the manufacturer's overnight protocol with minor modifications.CSF was collected once in a subset of ADNI participants at baseline. Aβ1–42, 2.2–6.3% for tau and the inter-run %CV for CSF pool samples ranged from 5.1 to 14% for Aβ1–42, 2.7–11.2% for tau.CSF was collected into polypropylene tubes or syringes provided to each site, and then be transferred into polypropylene transfer tubes without any centrifugation step followed by freezing on dry ice within 1 h after collection for subsequent shipment overnight to the ADNI Biomarker Core laboratory at the University of Pennsylvania Medical Center on dry ice. Aliquots (0.5 ml) were prepared from these samples after thawing (1 h) at room temperature and gentle mixing. The aliquots were stored in bar code-labelled polypropylene vials at −80 °C. Fresh, never before thawed, 0.5 ml aliquots for each subject's set of longitudinal time points were analysed on the same 96-well plate in the same analytical run for this study to minimize run to run and reagent kit lot sources of variation. Within run coefficient of variation (%CV) for duplicate samples ranged from 2.5 to 5.9% for AβApolipoprotein E (ApoE) and ferritin protein levels were determined using Rules Based Medicine .http://www.adni-info.org/).Further information on the procedures and standard operating procedures can be found in previous publications40www.loni.ucla.edu/ADNI) and for detailSubjects with a 1.5-T MRI and a sagittal volumetric 3D MPRAGE with variable resolution around the target of 1.2 mm isotropically were included in the analysis. See APOE genotype, and levels of CSF haemoglobin (Hb) and Factor H (FH). CSF Hb was included as a proxy for blood contamination, to control for the possibility of a traumatic tap introducing plasma ferritin into the CSF samples. FH was used to control for inflammation, since ferritin levels are known to be elevated in certain inflammatory conditions , linear mixed effects models were used. These models were adjusted for the same variables as the baseline models of cognition, and additionally included time as interacting variable with each of the CSF biomarkers. AD subjects were excluded from the longitudinal analysis because of low rate of follow up  and Rey Auditory Verbal Learning Test (RAVLT) scores with CSF ferritin, the CSF tau/Aβollow up . A signiAPOE-ɛ4 genotype as confounding variables together with CSF ApoE, tau/Aβ1–42 and ferritin. A minimal model containing only the CSF biomarkers was identified via BIC step down procedure and log likelihood test.Cox proportional hazards model was used to assess the impact of CSF analytes on the time to AD conversion. The initial model contained age at baseline, gender, years of education and 1–42 analytes were included in logistic regression models of MCI conversion to AD that were adjusted for age at baseline, gender, years of education, APOE genotype and BMI. These models determined the predictive performance of these analytes in identifying stable MCI participants from MCI participants who, up to 102 months, had a diagnosis change to AD. The receiver-operator curves and the area under the curve were derived from the predictive probabilities of the logistic regression models.Logistic regression analysis was used to assess the impact of CSF analytes on risk of conversion to AD. Combinations of CSF ferritin, ApoE and tau/Aβ1–42 with longitudinal structural (MRI) changes to hippocampus and lateral ventricle were analysed using linear mixed models adjusted for age, years of education, BMI, gender and APOE genotype and intracranial volume. For all models, CSF ferritin, ApoE, tau/Aβ1–42 and baseline diagnosis were included as fixed effects and were not removed from a minimal model. Two random effects were also included, intercepts and slope (time). An interaction between time and diagnosis, time and CSF ferritin, time and CSF ApoE, as well as time and CSF tau/Aβ1–42 were also included for all models. All the AD subjects were excluded from MRI analyses due to low numbers and short follow-up. PET imaging data from ADNI were not included in the analysis because there were too few patients who had CSF ferritin measured and who also underwent PET imaging at baseline.The relationships between CSF ferritin, ApoE, tau/AβHow to cite this article: Ayton, S. et al. Ferritin levels in the cerebrospinal fluid predict Alzheimer's disease outcomes and is regulated by APOE. Nat. Commun. 6:6760 doi: 10.1038/ncomms7760 (2015).Supplementary Figures 1-5 and Supplementary Tables 1-3


Donor cell derived malignancies are a rare and interesting complication of allogeneic bone marrow transplantation. We present a case of a 56-year-old male with donor cell myeloid sarcoma of the stomach and myocardium. Treatment of acute leukemia with an allogeneic bone marrow transplant provides a chance for a cure, although relapse is still a cause of mortality posttransplant. Most often when patients relapse, it is from the failure to eradicate their original leukemia. Rarely, patients may develop a leukemia derived from their donor cells, termed donor cell leukemia . We presA 56-year-old male was admitted to our hospital 704 days after a human leukocyte antigen- (HLA-) matched sibling donor (sister) allogeneic bone marrow transplant for myelodysplastic syndrome (MDS) with new abdominal pain, weight loss, and night sweats. His myelodysplastic syndrome was classified as refractory anemia with excess blasts-1. He had complex cytogenetics t, add (11)(p15), add (12)(p13), del (17)(p11.2),−22[17]/46,XY[3]) and positive fluorescence in situ hybridization (FISH) for 5q, 7q, and 11q23. Prior to this hospitalization, he was in complete remission with normal peripheral blood counts, normal trilineage hematopoiesis on bone marrow exam, and complete donor chimerism (>98%) of his peripheral blood.Computed-tomography scan of the chest and abdomen was performed and revealed gastric wall thickening , retrope2 per day on days 1–7 and daunorubicin 90 mg/m2 per day on days 1–3). Fourteen days after induction of chemotherapy, a positron emission tomography scan was done to evaluate the response to chemotherapy. The gastric wall had residual disease with a standard uptake value of 6.3 and the interventricular septum of the heart had increased uptake when compared to the rest of the myocardium. Subsequently, he was reinduced with HiDAC (high dose cytarabine 2000 mg/m2 twice a day for 3 days), but he developed neurotoxicity and then bone marrow relapse. He died 12 weeks later. At this time, the donor had a normal CBC.The patient received induction chemotherapy with “7 + 3”  is a rare complication of allogeneic transplantation. The first case was reported inw et al.  and a rew et al. , gingivaw et al. , and B-cw et al. . Donor-dw et al. . Then, 5DCL is also very rarely reported in recipients of solid organ transplants. In 1999, Bodó et al. reported a patient with donor-derived acute promyelocytic leukemia after a liver transplant . SubsequThe etiology of DCL still remains poorly understood. The most simplistic explanation is that the graft harbored an occult malignancy that is below the threshold of current screening at time of donation and over time develops into overt leukemia. This notion seems tenable in patients who develop donor cell leukemia and their donor develops a malignancy with the same phenotype , 9. HoweEstablishing the diagnosis of DCL has evolved significantly over the past 40 years paralleling the more frequent use of molecular techniques. Currently used methods of distinguishing between donor and recipient cells include XY FISH (in sex-mismatched donors) and variable number of tandem repeats (VNTR), as well as STR . In our The prognosis of DCL has been reported to be similar to patients with other secondary malignancies, and there are no prospective trials investigating outcomes in this patient population. Retrospective reviews of published case reports have found that, similar to primary AML, standard AML therapy in some patients leads to a complete response (CR) and some patients may benefit from a second stem-cell transplantation for long-term control. According to a review of 64 cases of DCL or MDS, 34 cases or 53% of patients died with a median survival of 5.5 months and only 17 or 26% had obtained a CR when the authors had reported their cases . While D


Macroscopic real-time evaluations of the histopathology and degree of invasion of colorectal polyps help to select the most suitable endoscopic treatment method. Dual-focus (DF) narrow-band imaging (NBI) is a new imaging enhancement system that uses digital and optical methods to enhance the view of blood vessels on mucosal surfaces. However, the superiority of this technique over standard imaging techniques has not been previously reported. The aim of this study was to determine whether the two-stage optical systems in a new generation of endoscopes will increase the diagnostic accuracy of colorectal polyp recognition.The study included 270 patients, and 386 colorectal polyps were diagnosed and removed. The polyps were assessed with white light and NBI using one- and two-stage optical systems, respectively. After being classified according to the Kudo pit pattern schemes, the polyps were removed and histopathologically verified.L lesions  and Kudo IIIS lesions . NBI-DF also increased the accuracy of preliminary diagnoses of Kudo IV lesions . The Kudo V pit patterns were also more distinct with NBI-DF imaging, increasing the diagnostic accuracy from 91.67 to 100 %.Regarding non-neoplastic lesions (Kudo I and II), no difference was observed in the recognition of polyps when using the NBI-DF function. We observed improved accuracy in the preliminary diagnoses of Kudo IIIUsing a two-stage optical system with electronic colorization of the mucosa increased diagnostic accuracy for differentiating colorectal polyps with neoplastic potential. Colorectal cancer (CRC) is the third most common cancer and the second leading cause of cancer deaths worldwide . Most coIn addition to NBI, a novel diagnostic technique has recently emerged, the so-called dual-focus (DF) magnification . DF two-stage optical lens technology from Olympus allows physicians to switch from a normal focus mode to the near focus (NF) mode, enabling up to 100× magnification with the single push of a button. This visualization allows for the close examination of mucosal tissue and capillary networks.The aim of this study was to evaluate and compare the diagnostic characteristics of a novel NBI system with DF magnification function in differentiating colorectal polyps.The study was conducted at MEDICINA Specialist Diagnostic & Therapeutic Centre , in Cracow, Poland. The study was approved by the local ethics committee and was conducted in accordance with the principles of the Declaration of Helsinki. As a clinical trial, the study was registered in a centralized clinical trials registry . All authors of this study had access to the study data and had reviewed and approved the final manuscript.We included patients 40–65 years old who underwent the procedure in the context of an opportunistic colorectal cancer screening. In addition to age, the following exclusion criteria were applied: symptoms of colon cancer , changes in bowel movement regularity, and unexplained weight loss. Patients who had already received a colonoscopy within the last 10 years were also excluded. A total of 2806 patients were referred and scheduled for outpatient colonoscopy in 2012 as a part of a national colorectal cancer-screening program. Of these patients, 842 were examined using a 190 series Exera III NBI system  with DF capability. The cecum was successfully intubated in 771 patients, whereas the remaining 71 subjects were excluded due to inadequate bowel preparation or neoplastic infiltration occluding the lumen. Colorectal polyps were detected in 270 patients (35 %) who were prospectively enrolled in the study  in 4 L of water taken in 4 doses every 6 h one day before the colonoscopy). Each colonoscopy was performed by one of the three experienced endoscopists (each of whom has performed more than 5000 colonoscopies).S, IIIL, or IV, as well as a subset of VI, are intramucosal neoplastic lesions . Lesions with a type VN pattern and a subset of type VI are suggestive of deep invasive carcinoma  was used for all procedures in this study. Moreover, all examinations were performed with magnetic endoscope imaging , which helped to avoid loop formation upon insertion and improved the accuracy of anatomical localization and endoscope positioning. These changes were crucial not only for conducting an effective procedure but also for localizing the pathological lesions more precisely within the large intestine. After cecal intubation, the colonic mucosa was carefully visualized with WL while withdrawing the colonoscope. All polyps detected during the procedure were documented for size, location, and morphology . Each polyp was routinely evaluated in real time, initially with WL and later with NBI and NBI-DF. All images were captured and stored as high-definition JPEG files . Histology was predicted for all polyps in vivo using WL, NBI, and NBI-DF based on the surface mucosal and vascular patterns identified by the respective techniques. Following these evaluations, the lesions were classified according to Kudo’s pit pattern classification and subsequently resected endoscopically or surgically. Kudo proposed a gross classification of pit patterns into seven types. It has been suggested that type I and II pit patterns are characteristic of non-neoplastic lesions, such as normal mucosa or hyperplastic polyps. However, most lesions showing pattern types IIIoma Fig. , 6, 7, 82 test. The Mann–Whitney U test was used to compare quantitative variables between two groups. The Kruskal–Wallis test was used for comparisons of quantitative data in more than two groups. The materiality threshold was established at p ≤ 0.05.The materials acquired in this study were systematized and analyzed, and a distribution of the variables was established. Because the analyzed parameters do not have a normal distribution, nonparametric tests were used in the analysis. Qualitative variables were compared using the independent χA total of 270 patients  were recruited. The number of patients in each age group is summarized in Table 7.41 % an2.59 %; mL, 28.24 %; Kudo IIIS, 30.57 %; and Kudo IV, 4.4 %); and 3.11 % of the polyps were invasive (Kudo V). Using NBI imaging together with the DF function (NBI-DF), we detected 35.2 % potentially non-neoplastic polyps , 36 % noninvasive , and 2.85 % invasive polyps (Kudo V) ; 63.2 % of the polyps were noninvasive (Kudo IIIL (87.16 %) and Kudo IIIS (87.29 %). High-grade dysplasia polyps (4.92 %) were described in vivo as Kudo IV (88.24 %). For malignant lesions, the mucosal pattern was usually designated as invasive pit pattern Kudo V (91.67 %)  or Kudo II (89.55 %) pit patterns, and low-grade adenomas were primarily predicted as Kudo IIIL 8.16 % andL and 92 % of Kudo IIIS polyps were low-grade adenomas; and 94.12 % polyps with noninvasive pit patterns (Kudo IV) were high-grade adenomas. All Kudo V polyps with an invasive pit pattern had a structure of adenocarcinoma upon histopathological examination  and the Kudo IIIS lesions . In the case of Kudo IV pit patterns, NBI-DF increased the accuracy of preliminary diagnoses from 88.24 to 94.12 % (p < 0.01). Kudo V pit patterns were also more distinct on NBI-DF imaging, improving from 91.67 to 100 % .Another concern is the recommended method of observing polyps measuring <5 mm localized in the distal part of the large intestine. Leaving these polyps in place lowers the procedure cost; however, this choice is not cost-effective, in part because of liability concerns. Not removing the polyps is less expensive, but the sequelae of diagnosing CRC in the location of previously unremoved polyps for observation can have fatal consequences. Using our methods, all polyps can be removed. According to the PIVI criteria, we gain >90 % conformity of the macroscopic image in NBI-DF upon histopathological examination, regardless of the polyp size . NeverthL or IV pits) are recognized at the margins of the resected tumor. In the future, NBI might contribute to real-time histological analysis during colonoscopy, which could substantially reduce the risk of polypectomy and the costs of histological evaluation by allowing adenomatous polyps to be resected and discarded.In conclusion, this study shows that NBI with optical magnification is highly accurate in predicting polyp histology in real time using a simple pattern classification system. Characteristic pit patterns obtained by magnifying NBI endoscopy provide useful clues regarding the differentiation of adenomatous from non-adenomatous polyps in vivo, without using dye. The pit pattern observations using magnifying NBI colonoscopy were also useful for assessing the resected margins after polypectomy or endoscopic mucosal resection. It may be necessary to perform subsequent management procedures, such as hot biopsies or argon plasma coagulation procedures, when neoplastic pit patterns (Kudo’s IIIUsed appropriately in experienced hands, this technique has potential as a valuable adjunct to standard colonoscopy in predicting the histological characteristics of colorectal polyps.


Pneumocephalus following ventriculoperitoneal (VP) shunt insertion is an exceptionally rare occurrence. We report such an event after attempting ventricular puncture (ventriculostomy) for VP shunt insertion and then discuss the management of the same. Dry tap can lead to multiple attempts for ventriculostomy with the associated added risks of complications, as well as complicating the subsequent management. In addition, there is an increased risk of tension pneumocephalus, seizure and shunt failure due to a blockage by air bubbles. Our patient presented with features of raised intracranial pressure two months following craniotomy and evacuation of traumatic subdural hematoma. External ventricular puncture revealed egress of CSF under pressure. Upon attempting VP shunting for post-traumatic hydrocephalus, we experienced dry tap during ventricular puncture that complicated further management. We placed the proximal shunt in the presumed location of the foramen of Monro of ipsilateral frontal horn of lateral ventricle and did not remove the external ventricular drain. Post-operative CT scan revealed pneumoventriculi as the cause for the dry tap during ventricular puncture. Patient was managed with 100% oxygen. He showed gradual improvement and was later discharged. This case shows that variations in the procedure, including head down positioning, adequate cruciate dural incision prior to cortex puncture, and avoiding excessive egress of CSF can help to prevent such complications. Tension pneumocephalus is a life-threatening emergency that necessitates immediate surgical intervention3. It is rarely reported after cerebrospinal fluid (CSF) diversion procedures5. We present a rare case of tension pneumocephalus, resulting in dry tap during ventriculostomy and discuss its subsequent management.Pneumocephalus is defined as the presence of air within the calvarium. It often follows trauma but is also a common sequelae of intracranial surgeryex vacuo due to volume loss and changes in CSF dynamics subsequent to the previous accident, external ventricular drainage (EVD) was placed which revealed egress of CSF under pressure. The reasons for opting to choose EVD prior to VP shunting are threefold. Firstly, we had to rule out post traumatic hydrocephalusex vacuo by measuring the opening pressure of the CSF egress and looking for the neurological improvement in the patient following CSF diversion. Secondly, since we not have the programmable VP shunting available, we need to appropriately choose the Chabra shunt depending on the opening pressure so as to prevent either over drainage or under drainage of CSF. Lastly, since it was a post traumatic case, we need to measure the CSF protein (as it may be increased from the lysed traumatic subarachnoid blood) and also we need to rule out subclinical meningitis. Both these may be the reasons for shunt failure. Following EVD, the patient showed gross improvement in his previous deficits. CSF sugar and protein was within range. Gram stain was negative for any bacteria. Repeat CT scan post EVD did not reveal any pneumocephalus. Thereafter he was scheduled for insertion of a VP shunt. EVD was clamped for 6 hours prior to the procedure to facilitate the ventricular tap. During insertion of the VP shunt, there was dry tap during an attempt of ventriculostomy from the Kocher’s point. We made two further attempts to ensure the correct trajectory of the shunt end and also to reflush the shunt end to prevent blockage due to blood clots and cell debris. We placed the shunt tip in the presumed location of the foramen of Monro of the frontal horn of ipsilateral lateral ventricle. We did not remove the EVD, hoping that it would act as a safety channel for CSF bypass had we missed the correct trajectory for the VP shunt.Herein we report a case of a 35-year-old male from Nawalparasi, Nepal, who had undergone a craniotomy and evacuation of acute subdural hematoma following an automobile accident 2 months before admission to our institution. He presented with complaints of an abnormal gait, with a tendency to fall backwards and also with features of frontal lobe-related incontinence. There were no significant past medical illnesses. He was taking Sodium Valproate  as seizure prophylaxis following the traumatic head injury and surgical intervention for the same 2 months previously. Fundus examination revealed the presence of papilledema. A head computerized tomography (CT) scan revealed the presence of evolving hydrocephalus. To rule out hydrocephalusth day post-operation showed that the proximal shunt was in the third ventricle  was also ensured.A postoperative scan revealed the presence of tension pneumocephalus and pneumoventriculi . The patntricle  and ther1. The incidence of this entity was reported to be as high as 100% following supratentorial craniotomies6. On the other hand, tension pneumocephalus is a neurosurgical emergency that requires rapid surgical intervention. Pneumocephalus as a complication of CSF diversion procedures is rare5. Diagnosis is mainly based on clinical examination and computerized tomography (CT) scan4. Two CT findings that characterise the condition – the ‘Mount Fuji’ sign and the ‘air bubble’ sign - have been described by Ishiwataet al.7.Pneumocephalus usually occurs after head trauma, skull base fractures, and associated CSF fistulas4. It is claimed that moderate cerebral atrophy might play a role1.There are two factors that are thought to be responsible for tension pneumocephalus development. The first is a decrease in intracranial pressure due to a sudden egress of CSF; the second is the presence of a craniodural defect that works as a one way valve allowing air inflow to the intracranial space and preventing outflowThe duration of the shunt surgery must be as short as possible and CSF leakage during the connection of the shunt system must be avoided. Another factor that can lead to undesirable outcomes can be introduced during the puncturing of the cortex. Adequate cruciate incision must be given to prevent the passage of environmental air into the subdural space. Filling the subdural space on the ventriculostomy site with irrigation fluid until overflowing might help the outflow of air from the intracranial vault, reducing the risk of this rare complication. Cortical atrophy may have also had an effect on isolated air collection within the subdural space. In our case, another remote possibility for the development of pneumocephalus would be any leak in the closed drainage system of the previous EVD drain. Properly layered closure of the skin in VP shunt surgery is the most important factor for prevention of this rare complication.The pneumocephalus in our case most likely would have occurred either during trephination of burr hole or while incising the dura prior to ventricular tap. Other probable reasons were ruled out since the EVD was clamped  and we also ensured the air locked system of the EVD circuit (as breach in the same would let the air get sucked in).The dry tap, as seen in our case, can lead to multiple attempts to attain the correct shunt trajectory, thereby increasing the risk of false trajectories and track hematomas. If there had been no EVD, then this would have led to termination of the procedure, thereby adding to the morbidity and risk of subsequent surgery. One alternative to our approach would be the use of intra-operative CT scan to ensure the diagnosis. Unfortunately this is not currently possible in developing countries like ours. We can, if available, take help of neuro-navigation tools to ensure the correct trajectory to the ventricles even in cases were in pneumocephalus occurs. This can reduce the added burden of subsequent surgeries and associated risk of anesthesia.et al.8 have advocated the use of 100% oxygen for rapid resolution of pneumocephalus.There is also risk of seizure and rapid neurological deterioration due to tension pneumocephalus. Once this occurs, close monitoring of the patient, rapid and accurate identification of tension pneumocephalus, and immediate surgical intervention is life-saving. GoreIn conclusion, though VP shunting is one of the most common surgical procedures performed in neurosurgery, strict adherence to basic principles should be followed during the procedure so as to prevent avoidable complications such as in our case which may in times lead to sudden deterioration in the patient and also add to diagnostic and therapeutic dilemma to the concerned surgeons. One advantage in this case was the presence of an EVD as a safety bypass for CSF diversion. The limitations of our approach can be attributed to the unavailability of intra-operative CT scan and neuronavigation techniques which would have aided in early diagnosis and management in this scenario.Both written and verbal informed consent for publication of images and clinical data related to this case was sought and obtained from the wife of the patient. The authors provide important guidelines for the evaluation and management of hydrocephalus developing in the post-craniotomy follow-up period after surgical traumatic acute subdural evacuation. The authors presented an interesting case, which presented with gait abnormality associated with falling back tendency and underwent craniotomy surgery two months earlier for evacuation acute subdural hematoma. A cranial computed tomography revealed presence of hydrocephalus. In the standard management of post traumatic craniotomy development of hydrocephalus, it is essential to evaluate the possible causes, degree of intracranial pressure rise, evaluating beneficial effect of cerebrospinal fluid diversion procedure i.e. ventriculoperitoneal shunt or thecoperitoneal shunt, assessment of CSF protein level, and exclusion of associated indolent meningitis as possible causes.1Ideally for excluding ex-vacuo post-traumatic hydrocephalus development, CSF sampling can be done through lumbar drainage or by placing external ventricular drainage and assessing initial CSF egress pressure with manometry for selection of pressure range and most suitable type of future shunt requirement, cytological, protein level for suitability of shunt functioning after shunt insertion; as very high protein level can be relative contraindication for shunt, biochemical study and culture sensitivity studies to exclude infective pathology and assessing Mini mental scale for cognitive assessment and gait changes . The patient shows improvement following CSF drainage CSF diversion surgery in the form of thecoperitoneal or ventriculoperitoneal shunt, however the former is the preferred choice.2. After air entering the cranial cavity leads to rise in the intracranial pressure3.   Development of pneumocephalus requires communication between arachnoid membrane and associated defect in the dura, calvarium and communication to the external environment. As CSF egresses, air enters the intracranial cavity and development of pneumocephalus. The “ball valve” mechanism was postulated by Dandy, as air travels in only one direction4. The author postulated negative intracranial pressure resulting from excessive loss of CSF through an iatrogenic lumbar drain or settling into the distendable spinal subarachnoid space or simply drainage via normal pathways with physiologic activity such as inspiration or the Valsalva manoeuvre4. However, when there are fistulous communications between the intracranial and outer environment, air can enter the intracranial space as a consequence of negative pressure gradient3,5-6. As the authors' case had EVD, that might have unfortunately disconnected and leak of CSF led to development of tension pneumocephalus. In cases of dry tap, insertion of the ventricular end is specially challenging and options include abandoning the procedure,  getting an intraoperative cranial CT scan or help of an endoscope, or image guidance or infusing fluid through the EVD into the ventricle thereby distending the ventricle prior to reattempting Ventricular catheter insertion. However, dry tap is really a neurosurgical emergency needing tactful management, which depends on available facility at surgical centre including image guidance, intraoperative CT scan facility, endoscope facility and expertise of surgical team.The other postulates for tension pneumocephalus is the “Inverted-soda-bottle effect”, proposed by HorowitzWe have read this submission. We believe that we have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. The authors analysed dry tap during ventriculostomy in the presence of EVD inserted few days before.Their explanations are reasonable. But there might be additional explanations.The EVD was inserted to rule out ex vacuo hydrocephalus.  CSF egressed under pressure. Here the authors could insert VP shunt on first occasion (without previous EDV).   There is no report on patient's condition between EVD and VP insertion.  If it improved and remained stable, no significant tension hydrocephalus is expected to evolve. It is my theory that the air was sucked into the ventricles and trapped subdurally at the time of VP shunt insertion when EDV was probably not stopped enough long time before VP insertion.   EDV produced some minor negative pressure.  During attempts of inserting ventricular catheter of VP shunt produced sucking of the air into the ipsilateral ventricle and trapping it there.  Similarly, during trephination of a burr hole for insertion of VP shunt  and cutting the dura, some air was sucked subdurally on that side.   As slowly evolving postraumatic  hydrocephalus is mainly of malabsorptive communicative type, lumbar drainage is less invasive attempt to reveal ex vacuo hydrocephalus than EDV. EDV or LD is stopped cca. 4-6h before inserting VP catheter to reexpand the ventricles and ease targeting.I have read this submission. I believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. Respected sir,Thank you for your report.We place the EVD in our ICU in suspected cases of hydrocephalus and  then plan for the shunting if there is egress of CSF under pressure and neurological improvement in the patient. This case showed neurological improvement after EVD placement. The repeat CT after the shunt did not reveal pneumocephalus. We routinely do clamp the EVD 6 hours prior to the procedure to facilitate the tap.As you have correctly mentioned ,the pneumocephalus might have resulted either during the trephination or during corticol incision prior to ventricular tap.Sorry to have missed these points in our paper. VP shunt is the most frequent surgical technique in neurosurgery to treat hydrocephalus related with trauma, SAH, tumor and other brain diseases. Dry tap, which is a rare episode during the shunt operation, may cause confused decision and leading to complications. In this report, authors described the details of the dry tap and analysis the possible reasons behind, which is helpful for neurosurgeons to prevent this rare complication in clinical settings.I have read this submission. I believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.


Cardiac allograft vasculopathy (CAV) is one of the main causes of late-stage heart failure after heart transplantation. CAV is characterized by concentric luminal narrowing of the coronary arteries, but the exact pathogenesis of CAV is still not unraveled. Many researchers show evidence of an allogeneic immune response of the recipient, whereas others show contrasting results in which donor-derived cells induce an immune response against the graft. In addition, fibrosis of the neo-intima can be induced by recipient-derived circulating cells or donor-derived cells. In this review, both donor and recipient sides of the story are described to obtain better insight in the pathogenesis of CAV. Dual outcomes were found regarding the contribution of donor and recipient cells in the initiation of the immune response and the development of fibrosis during CAV. Future research could focus more on the potential synergistic interaction of donor and recipient cells leading to CAV. Cardiac transplantation is often successfully applied in the treatment of end-stage heart failure . Since 1One of the main causes of chronic rejection is cardiac allograft vasculopathy (CAV) –5. CAV iCAV is characterized by diffuse intimal thickening leading to progressive narrowing of the coronary arteries , 11. TheThe exact mechanism in which CAV is induced after heart transplantation is not elucidated, but it is known that both donor and recipient cells are involved . The queAccording to multiple research groups, the onset of CAV is caused by an immune response of the recipient against the donor , 9, 18. (1) In the direct pathway, recipient T-lymphocytes are activated after recognition of allogeneic MHCs (with a foreign antigen) of donor antigen presenting cells (APCs) .(2) The indirect pathway is activated by allo-recognition of processed foreign antigens by APCs of the recipient itself . The recIn all three pathways, the activation of T-lymphocytes will lead to secretion of cytokines such as interleukin-2 (IL-2) and interferon-γ (IFN-γ) . CytotoxIn addition, there is also evidence that “autoimmunity” plays an important role in the development of CAV . For exaFurthermore, the development of anti MHC-class-1-chain-related-A (MICA) antibodies may play a role in the development of CAV , 25. NorCytomegalovirus (CMV) infection has also been known to affect transplantation outcome and CAV. Systemic replication of CMV is associated with increased risk of rejection of the graft and the development of CAV . The theThe trigger of these responses  ultimately leads to the proliferation of smooth muscle cells (SMCs), accumulation of extracellular matrix and hyperplasia of the intima of the vessel wall Fig.  15, 22,,22, 23.Next to the recipient-derived immune response, there is also evidence that donor factors are involved in the immune-pathogenesis of chronic rejection and CAV , 35. DonDonor-derived cells also play an important role in the pathogenesis of CAV. The current theory is that remaining donor cells within the transplanted heart are able to actively induce an immune response of recipient immune cells , 42. It In addition to donor DCs, the functional activity of donor CD4 T-lymphocytes was studied. In a mouse model, the development of autoimmune reactions after heart transplantation and the contribution to CAV was analyzed . Donor CThe expression of donor programmed death-ligand 1 (PD-L1) is also involved in the development of CAV . This liIn addition, it has been shown that donor-derived selectins play an important role in the development of CAV . SelectiAccording to this accumulating evidence, there is a donor-derived immune response causing the development of allograft vasculopathy and donor cells are involved in regulating the allo-immune response of the recipient. Since this is a relatively new insight, more focus on these aspects is needed to reveal the exact mechanism and to define all of the donor and recipient cells involved.Interesting differences in transplantation outcome have been reported between males receiving a female heart or females receiving a male heart, the so called donor-recipient gender mismatch transplantations . Donor-rAt the organ level, contrasting studies showed higher incidents of rejection of female recipients receiving a male heart . These hBased on these findings, donor-recipient gender mismatch in heart transplantation is followed by dual outcomes. Transplantation of male hearts into females is characterized by higher rejection rates, but in the end, a higher long-term survival . TransplThe before-mentioned immune reactions could be followed by fibrosis, which plays an important role in the progressive thickening of the neo-intima and subsequently in the development of CAV . In some+CD34+Flk1 [VEGF-R1]+ in bone marrow, CD31 [PECAM-1]+CD146+vWF+NOS+ in circulation), are bone marrow-derived cells, which have endothelial regenerative properties [There is evidence emerging for the role of recipient endothelial progenitor cells (EPCs) in CAV . EPCs  and epithelial-to-mesenchymal transition (EMT) have shown to be potential contributors to neo-intima formation –78. EndoNext to individual donor-derived cells, it has been suggested that atherosclerotic plaques in coronary vessels of the donor, pre-existing in the transplanted heart, influence the outcome of CAV in the recipient . The athIn conclusion, it appears that not only inflammatory cells of the donor are involved in the pathogenesis of CAV, but also donor-derived cells, such as endothelial cells, epicardial cells, and smooth muscle cells. In addition, presence of atherosclerotic plaques might be correlated to fibrotic lesions in CAV. These findings provide new insights in a possible role for the donor in the development of neo-intima fibrosis in CAV; however, this is still under debate.Activation of the immune system of the recipient, for example by contact of donor- and recipient cells, should be avoided. Ideally, when transplanting a solid organ, there is no transfer of donor immune cells to trigger an immune response. In the future, this might be achieved by, for instance, ex vivo perfusion of the donor organ, clearing all immune cells from the graft . When exMultiple prophylactic approaches have been used to prevent the development of CAV . ModificCurrently used therapeutic interventions, such as statins, angiotensin-converting enzyme (ACE) inhibitors and immunosuppressive medications are not always successful in the treatment of CAV . These tCAV is the leading cause of death after heart transplantation, although the pathogenesis is not fully resolved . Until rIn conclusion, CAV is a complex disease with an unrevealed pathogenesis. Presumably, CAV is not induced only by the donor or recipient. Based on current research, it is clear that both donor and recipient cells are involved , 42, 43.


Normalized mean transit time (TT) in the left atrium has previously been shown to approximate left ventricular end diastolic pressure (LVEDP). In the present study, we characterized global and segmental central circulatory TT, in patients with and without systolic dysfunction, and delineated the relationship of TT to hemodynamics.Forty nine subjects undergoing clinically indicated right and left heart cardiac catheterization were prospectively recruited to undergo MRI in a 1.5T scanner. First pass perfusion using steady state free precession saturation recovery sequence was performed with gadolinium infusion at 0.01 mmol/kg. Global TT was defined as the time between the peaks of time intensity curves between the right atrium and the ascending aorta. Segmental TT included TT between right atrium to pulmonary artery (right heart TT), pulmonary artery to left atrium (pulmonary TT), or left atrium to ascending aorta (left heart TT). All TTs were normalized to heart rate. Multivariate regression analysis was performed to delineate the relationship of hemodynamic parameters measured during cardiac catheterization to TT. Receiver operating characteristic (ROC) analyses were performed to assess ability of global and segmental TT to predict elevated LVEDP.Characteristics of the study group included average age of 62 years, LV ejection fraction (EF) 47±16%, and LVEDP 18 ± 9 mmHg. Global, pulmonary and left heart TTs were significantly prolonged in patients with elevated LVEDP compared to patients with LVEDP<15 mm Hg. Of the TTs assessed, global and pulmonary TT were strongly associated with hemodynamic abnormalities related to left heart failure in univariate analysis. In multivariate regression analysis adjusting right atrial pressure, pulmonary artery (PA) peak systolic and mean pressure, pulmonary vascular resistance, LVEDP and PA oxygen saturation, global TT was most significantly associated with LVEDP (p<0.001) whereas pulmonary TT was most significantly associated with LVEDP (p=0.009) and PA oxygen saturation (p=0.044). Using the ROC analysis, prolonged global TT and pulmonary TT were predictive of elevated LVEDP with an area under the curve (AUC) of 0.79 and 0.77, respectively. A cutpoint value of global TT ≥10.5 cardiac cycles and pulmonary TT ≥ 8.4 cardiac cycles were predictive of elevated LVEDP with sensitivity of 82% , 54% and specificity of 71% or 100%, respectively.Prolonged global and pulmonary TT were strongly associated with elevated LVEDP in patients with and without systolic dysfunction. Our findings suggest that global and pulmonary TT evaluation is promising in assessment of LVEDP elevation by MRI.N/A.