Data from several rat strains/lines strongly suggest that catalase-mediated brain oxidation of ethanol into acetaldehyde is an absolute requirement (up 80%–95%) for rats to display ethanol’s reinforcing effects and to initiate chronic ethanol intake. Acetaldehyde binds non-enzymatically to dopamine forming salsolinol, a compound that is self-administered. In UChB rats, salsolinol: (a) generates marked sensitization to the motivational effects of ethanol; and (b) strongly promotes binge-like drinking. The specificity of salsolinol actions is shown by the finding that only the R-salsolinol enantiomer but not S-salsolinol accounted for the latter effects. Inhibition of brain acetaldehyde synthesis does not influence the maintenance of chronic ethanol intake. However, a prolonged ethanol withdrawal partly returns the requirement for acetaldehyde synthesis/levels both on chronic ethanol intake and on alcohol relapse-like drinking. Chronic ethanol intake, involving the action of lipopolysaccharide diffusing from the gut, and likely oxygen radical generated upon catechol/salsolinol oxidation, leads to oxidative stress and neuro-inflammation, known to potentiate each other. Data show that the administration of N-acetyl cysteine (NAC) a strong antioxidant inhibits chronic ethanol maintenance by 60%–70%, without inhibiting its initial intake. Intra-cerebroventricular administration of mesenchymal stem cells (MSCs), known to release anti-inflammatory cytokines, to elevate superoxide dismutase levels and to reverse ethanol-induced hippocampal injury and cognitive deficits, also inhibited chronic ethanol maintenance; further, relapse-like ethanol drinking was inhibited up to 85% for 40 days following intracerebral stem cell administration. Thus: (i) ethanol must be metabolized intracerebrally into acetaldehyde, and further into salsolinol, which appear responsible for promoting the acquisition of the early reinforcing effects of ethanol; (ii) acetaldehyde is not responsible for the maintenance of chronic ethanol intake, while other mechanisms are indicated; (iii) the systemic administration of NAC, a strong antioxidant markedly inhibits the maintenance of chronic ethanol intake; and (iv) the intra-cerebroventricular administration of anti-inflammatory and antioxidant MSCs inhibit both the maintenance of chronic ethanol intake and relapse-like drinking.This review article addresses the biological factors that influence: (i) the  The mechanisms leading to alcohol rejection in the virtually abstainer UChA line are mainly related to polymorphisms in nuclear and mitochondrial genes that lead to a slow metabolism of acetaldehyde and to high blood acetaldehyde levels. These UChA studies have been previously described  of crosses between high-intake (C57BL) and low intake animals (DBA) results in animals presenting the complete alcohol intake phenotype spectrum spanning their original strains .A number of studies in laboratories in Spain, Chile and Italy, using Sprague-Dawley, UChB or Wistar rats, respectively  will bar-press to self-administer both ethanol and acetaldehyde into the ventral tegmental area (VTA). Noteworthy, the levels of acetaldehyde required for self-administration into the VTA are three orders of magnitude lower for acetaldehyde, in the range of 10 μM than those for ethanol, which are in the range of 10–20 mM Genetic inhibition of catalase synthesis. Figure (b)Transducing a gene encoding an enzyme that degrades acetaldehyde. The administration to naïve UChB rats of a lentiviral vector coding for the wildtype high affinity Aldh2, aimed at increasing the VTA ability to degrade acetaldehyde resulted in an 85% inhibition of ethanol (10% v/v) intake , an enzyme that generates acetaldehyde, acquisition (development) of ethanol reinforcement in rats.The above studies would preclude other brain systems in the ALDH2*2 genotype). However high systemic concentrations of acetaldehyde prior reached when acetaldehyde itself is orally consumed or it is administered intraperitoneally which can cross the blood brain barrier and are reinforcing, as shown by Peana et al. -salsolinol -SAL at concentrations of 0.03–0.3 μM is self-administered intra VTA by Wistar rats. These concentrations of (R/S)-SAL are one to two orders of magnitude lower than the concentrations required for acetaldehyde self-administration in the same brain area.Ethanol-derived acetaldehyde condenses non-enzymatically with brain dopamine to generate racemic (R/S)-SAL into the VTA of Wistar rats also result in an increased release of dopamine in the nucleus accumbens  is endogenous dopamine required to generate SAL? (ii) does the chronic administration of (R/S)-SAL generate a sensitized state similar to that generated by chronic ethanol intake, which augments ethanol reinforcement? (iii) does (R/S)-SAL administration to naïve rats induce ethanol motivational effects? (iv) is there an enantiomer specificity distinguishing the effects of R-salsolinol vs. S-salsolinol; and (v) does (R/S)-SAL administration result in locomotor sensitization, as it happens after chronic ethanol administration?(i)In vitro studies by Melis et al. R/S)-SAL .Intra-cerebroventricular Figure  or systeL Figure  to ethanL Figure . This efL Figure  in Sprag(iii)R/S)-SAL increased the motivational effects of ethanol as shown by the place preference technique R/S)-SAL is also seen with R-SAL while the S-SAL enantiomer is inactive -SAL remain unclear, the specific effect of the R-enantiomer in inducing the motivational effects of ethanol, suggests that in vivo the chirality of the C-1 center of (R/S)-SAL plays an important role in changing its affinity for transporters or receptors associated with ethanol intake.Intracerebral administration studies in UChB rats showed that the ethanol motivational and intake sensitization effects of The administration of (R/S)-SAL to Wistar or UChB rats induced a sensitization to its locomotor effects .The failure of the anticatalase or ALDH2 coding lentiviral vectors to reduce ethanol intake in rats that had consumed alcohol for 2 months suggests that following chronic alcohol consumption other signaling pathways might be recruited. Chronic consumption of drugs of abuse induces changes at the molecular, cellular and neurocircuitry levels that mediate the transition from occasional, controlled substance use to loss of control in drug intake and chronic addiction  transporter (GLT1) in astrocytes of tripartite glutamatergic synapses. Sari et al.  learned cues  of many addictive drugs including: (i) cocaine  in the brain: (i) cocaine inhibits both the dopamine transporter and the norepinephrine transporter . Animals were chronically self-administering alcohol; thus not in an abstinent condition  are known to potentiate each other.Overall, the studies reviewed indicate that different mechanisms are responsible for the In UChB rats ingesting ethanol chronically for 2-months, a 4-week ethanol deprivation leads to a partial recovery of the inhibitory effect of the anti-catalase vector on ethanol intake Figure . The facThe relapse-like drinking also known as the “alcohol deprivation effect” (ADE) is a condition in which animals subjected to chronic ethanol intake followed by a long deprivation, consume intoxicating amounts of ethanol in as little as 60-min upon ethanol re-access. The ADE paradigm in animals has good predictive value in representing relapse-like drinking in humans as it is inhibited by three medications used clinically to reduce ethanol intake, namely: naltrexone, nalmefene and acamprosate-Ca  partly reduced ADE ethanol intake upon re-access.In the ADE model in UChB rats, chronic ethanol intake for 1–3 months is interrupted by an alcohol deprivation of 7–15 days before animals are allowed ethanol re-access. Upon ethanol re-access, animals consume intoxicating amounts of ethanol of the order of 2–2.5 g ethanol/kg/in the first 60 min of re-access  and possibly relapse-like drinking. Developments in the stem cell field have shown that most tissues contain mesenchymal stem cells . Figure 5 cells in 5 μL) injected into the brain lateral ventricle reduced by 40%–60% the maintenance of chronic alcohol intake for the 10 days studied.Recent work in UChB rats acquisition) chronic ethanol intake. In the UChB rat, brain-derived acetaldehyde was shown to be an absolute requirement (80%–95%) for the initiation of chronic ethanol intake.Confirm, by the use of genetic modifications, studies by several groups in several rat strains that had indicated that catalase-mediated brain oxidation of ethanol into acetaldehyde is required for animals to initiate ((b)maintenance) has been attained, brain acetaldehyde generation is no longer required to perpetuate its intake. This effect seen in UChB and Wistar rats has also been observed in operant ethanol self-administration studies in mice. Following a protracted ethanol deprivation, acetaldehyde is again required to induce a relapse-like condition.Demonstrate that after a steady chronic ethanol intake ((c)Demonstrate that the daily administration of the antioxidant drug N-acetyl cysteine to UChB rats that have consumed ethanol chronically markedly inhibits (70%–75%) voluntary ethanol intake. Noteworthy, N-acetyl cysteine did not inhibit the initial acquisition of ethanol intake of naïve animals.(d)Demonstrate in UChB rats that the intracerebral administration of salsolinol—the condensation product of dopamine and acetaldehyde—results in enhanced ethanol reinforcement, leading to binge-like ethanol intakes  and in an enhancement of ethanol motivational effects as shown by the place preference technique.(e)relapse-like ethanol drinking by 60%–85% for 40 days.Demonstrate that a single intra-cerebroventricular administration to UChB rats of mesenchymal stem cells, known to have marked anti-inflammatory and antioxidant properties, inhibited (f)acquisition of ethanol intake; while these metabolites are indirectly involved in maintenance of chronic ethanol intake and in relapse-like ethanol drinking. A new element playing a role in maintenance and relapse-like drinking is neuroinflammation, partly mediated by acetaldehyde increasing the diffusion of gut lipopolysaccharide into the systemic circulation and possibly by oxygen radicals generated in the oxidation of salsolinol.Overall, studies indicate that ethanol-derived metabolites are by themselves involved in the The studies presented:A final note, while many of the studies discussed have been conducted in rodents of different species and strains or in cells, extrapolation of these findings to humans requires caution.All the authors contributed to the writing and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


It is now increasingly appreciated that glial cells play a critical role in the regulation of iron homeostasis. Impairment of these properties might lead to dysfunction of iron metabolism and neurodegeneration of neurons. We have previously shown that dysfunction of glia could cause iron deposit and enhance iron-induced degeneration of dopamine (DA) neurons in Parkinson’s disease (PD). There also has been a substantial growth of knowledge regarding the iron metabolism of glia and their effects on iron accumulation and degeneration of DA neurons in PD in recent years. Here, we attempt to describe the role of iron metabolism of glia and the effect of glia on iron accumulation and degeneration of DA neurons in the substantia nigra of PD. This could provide evidence to reveal the mechanisms underlying nigral iron accumulation of DA neurons in PD and provide the basis for discovering new potential therapeutic targets for PD. Neuropathological hallmarks of PD include the degeneration and loss of dopaminergic neurons in the substantia nigra (SN) and the subsequent dopamine (DA) depletion in the striatum. Although the exact pathogenesis of PD is not fully understood, a growing body of research has confirmed that nigral iron accumulation was involved in the death of DA neurons in PD . Iron lepatients . In addipatients .Conventional MRI and diffusion-weighted imaging at 1.5 T have been recommended by European Federation of Neurological Societies (EFNS) to support a diagnosis of multiple system atrophy (MSA) or progressive supranuclear palsy versus PD . EFNS haTable 1. Our previous study and others have confirmed that increased iron levels were associated with increased expression of iron importer divalent metal transporter 1 (DMT1) and decreased expression of iron exporter ferroportin1 (FPN1) in PD animal and cell models . Increased iron and DMT1 expression were also observed in post-mortem PD patients  . There al models . The actl models  . In addition, cultured neurons, astrocytes and microglia all have the ability to store huge amounts of iron, but compared to neurons, glia can stored iron more effectively . In addition, studies found that iron overload could activate microglia and astrocytes and promote the release inflammatory factor and neurotrophic factors, which were involved in the regulation of iron metabolism of DA neurons  in microglia, thus producing many immune inflammatory factors  and tumor necrosis factor-α (TNF-α) , 1996. E factors . They fuFigure 3).In addition, our previous study showed that iron status of microglia can also affect secretion of pro-inflammatory cytokines including IL-1β and TNF-α and then participate in the degeneration of DA neurons . Our resFigure 3). A recent result showed that activated microglia could also stimulate astrocytes to release hepcidin via IL-6 signaling, which then prevented FPN1-mediated iron release and induced apoptosis of neurons . In addition, ceruloplasmin (Cp) is one of the major copper-binding proteins responsible for converting toxic ferrous iron into ferric iron (Another possible mechanism underlying the effect of microglia on iron metabolism of neurons is associated with interleukin-6 (IL-6). Studies have found that LPS could induce the expression and release of IL-6 in microglia. Released IL-6 from microglia could up-regulate hepcidin via IL-6/signal transducer and activator of transcription 3 (STAT3) signaling pathway in neurons . Hepcidi neurons   by protecting mitochondria, increasing the expression of copper and zinc-containing superoxide dismutase (Cu/Zn-SOD) and B-cell lymphoma-2 (Bcl-2) .In addition to release TNF-α, IL-1β, activated microglia can also release lactoferrin (Lf), which is an iron-binding protein belonging to the transferrin family. Lf-Lf receptor (LfR) transfers Fethan TfR . It was than TfR . In the than TfR . The expthan TfR . Immunohthan TfR . These fthan TfR . In VM n (Bcl-2) . This in (Bcl-2)  -induced neurotoxicity . They mesynuclein in astrocytes, showed that excess A53T α-synuclein in astrocytes caused severe astrogliosis, leading to dysfunction of astrocytes to maintain the integrity of BBB and homeostasis of extracellular glutamate. This induced inflammation, microglial activation and a significant loss of DA neurons in the midbrain in these mutant mice . Iron absorption mediated by these transporters in astrocytes can buffer high levels of extracellular iron, and thus inhibit high iron-induced damage to DA neurons. In addition, astrocytes can store iron via ferritin efficiently and release iron by FPN1. Cp is a kind of ferrous oxidase mainly in the form of glycosyl-phosphatidylinositol (GPI)-Cp in the brain . FPN1 and GPI-Cp co-expressed in the cell surface of astrocytes .Astrocytes participate in the formation of BBB and are generally accepted as principle contributor for the uptake of a variety of nutrients including iron to the brain. It controls the process of iron transport from outside the brain to inside the brain and regulates iron transport from astrocytes to other brain cells . Studies or TfR1 . Howeverortation . Most stin vitro . DMT1 isin vitro . DMT1 wain vitro  and mainin vitro , indicatin vitro  and resiin vitro  in astrohe brain  and can trocytes  to mediaIt has been reported that iron levels can affect the expression of iron related proteins in astrocytes. Ferric iron incubation increased ferritin expression and reduced the expression of TfR . This adFigure 4). These results confirmed that astrocytes can affect iron metabolism and survival of neurons by releasing neurotrophic factors BDNF and GDNF.Astrocytes may affect the iron metabolism of DA neurons in PD models. Astrocytes are vital for the survival of DA neurons by secreting various neurotrophic factors, such as brain derived neurotrophic factor (BDNF) and GDNF. Figures 4). This provides new experimental evidence on relationship between abnormal iron metabolism and inflammatory in PD. Further studies should be conducted to elucidate the exact mechanisms underlying the effect of LCN2 on iron accumulation in DA neurons in PD.Recently, another novel mechanism underlying neuron-glia interaction in iron metabolism has been reported . LipocalFigure 2). It has been shown that iron uptake of oligodendrocytes was accompanied by myelin formation and iron deficiency animals showed damage of myelin formation . It has been demonstrated that microglia-released ferritin is an important source of iron for oligodendrocytes -positive progenitor cells and differentiation into mature oligodendrocytes . Studies have shown that there were ferritin binding sites in oligodendrocytes . Another study showed that scavenger receptor class 5 (scara5) was the receptor of light chain ferritin (L-ferritin) expressed in embryonic mice and kidney cells of adult mice. Scara5 can bind to L-ferritin or Tf (but not HFt) and mediate its endocytosis  and reviewed their possible roles in the degeneration of DA neurons in PD  in this review. Glia could affect iron metabolism and survival of DA neurons in PD through the release of proinflammatory factors or neurotrophic factors. Excessive activation of glia aggravated iron accumulation and degeneration of DA neurons in PD. In addition, there exists a complex regulatory mechanism between glia, which leads to the final degeneration of DA neurons. Therefore, regulating the function of glia may provide a new therapeutic target for the treatment of iron-mediated neurodegenerative disorders especially PD.In recent years, considerable advances have been made in understanding iron metabolism in glia and neurons. We have summarized iron metabolism in glia (However, the current researches on iron metabolism rely mainly on experimental animals, especially in rodent models. These models cannot fully simulate changes of iron metabolism in the brain of PD patients. Therefore, it is crucial for studies using human stem cells, human glia or post-mortem brain tissue of PD patients to clarify iron metabolism of glia and their role in the degeneration of DA neurons in PD. Further investigations are also required to investigate whether the newly discovered transporters and regulatory proteins are also expressed in glia and how they functioned. In addition, the exact molecular and cellular mechanisms underlying the interaction between glia and DA neurons on iron metabolism should also be elucidated in the future.HX wrote the manuscript. YW and HX constructed the figures. NS and JW contributed to the editing of the manuscript. HJ and JX revised the manuscript. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


We have categorized the dataset from content and emission of terpene volatiles of peel and juice in both Navelina and Pineapple sweet orange cultivars in which D-limonene was either up- (S), down-regulated (AS) or non-altered  )  Specifications TableValue of the data•Volatile identification and quantification by HS-SPME and GC–MS can be categorized by Principal Component Analysis (PCA), which is helpful in the case of analyzing different and complex profiles to map out general trends in presence, accumulation and emission of specific chemical groups •We analyzed the terpene volatiles of peel and juice in both Navelina and Pineapple sweet orange cultivars with either up-, down-regulated or unaltered levels of D-limonene and related compounds. PCA can be a useful tool for rapid differentiation of fruit odors based on the comparison of volatile compound profiles •The statistic aggrupation of these specific or chemical groups of volatiles is helpful in defining which ones are the most influential for odor in each transgenic line.1Principal component analysis (PCA) revealed two major clustering groups in Navelina flavedo and juice with pulp in both analyses from individual volatiles or from groups of compounds: the down-regulated D-limonene fruits (AS3 and AS5) vs. the non-altered control fruits (EV) , Fig. 2.2GC–MS and SPME-GC/MS data of the volatile content in the transgenic and control orange fruits


The SAKK 19/05 trial investigated the safety and efficacy of the combined targeted therapy bevacizumab and erlotinib (BE) in unselected patients with advanced non-squamous non-small cell lung cancer (NSCLC). Although activating EGFR mutations were the strongest predictors of the response to BE, some patients not harboring driver mutations could benefit from the combined therapy. The identification of predictive biomarkers before or short after initiation of therapy is therefore paramount for proper patient selection, especially among EGFR wild-types. The first aim of this study was to investigate the early change in blood gene expression in unselected patients with advanced non-squamous NSCLC treated by BE. The second aim was to assess the predictive value of blood gene expression levels at baseline and 24h after BE therapy.Blood samples from 43 advanced non-squamous NSCLC patients taken at baseline and 24h after initiation of therapy were profiled using Affymetrix’ exon arrays. The 24h gene dysregulation was investigated in the light of gene functional annotations using gene set enrichment analysis. The predictive value of blood gene expression levels was assessed and validated using an independent dataset.Significant gene dysregulations associated with the 24h-effect of BE were detected from blood-based whole-genome profiling. BE had a direct effect on “Pathways in cancer”, by significantly down-regulating genes involved in cytokine–cytokine receptor interaction, MAPK signaling pathway and mTOR signaling pathway. These pathways contribute to phenomena of evasion of apoptosis, proliferation and sustained angiogenesis. Other signaling pathways specifically reflecting the mechanisms of action of erlotinib and the anti-angiogenesis effect of bevacizumab were activated. The magnitude of change of the most dysregulated genes at 24h did not have a predictive value regarding the patients’ response to BE. However, predictive markers were identified from the gene expression levels at 24h regarding time to progression under BE.The 24h-effect of the combined targeted therapy BE could be accurately monitored in advanced non-squamous NSCLC blood samples using whole-genome exon arrays. Putative predictive markers at 24h could reflect patients’ response to BE after adjusting for their mutational status.Trial registration ClinicalTrials.gov: NCT00354549The online version of this article (doi:10.1186/s12967-017-1174-z) contains supplementary material, which is available to authorized users. Combined targeted therapies represent novel therapeutic approaches simultaneously acting on several specific molecular pathways in cancer and having a number of advantages over standard single-targeted agents , 2.Several trials have shown the beneficial effect of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) in advanced non-small cell lung cancer patients (NSCLC) harboring activating EGFR mutations leading to the adoption of EGFR-TKI as standard treatment in this population , 4. PrecIn unselected patients, BE had better PFS than E alone without improvement of survival in recurrent NSCLC suggesting moderate activity of BE . As firsAlthough the presence of EGFR mutations is the strongest predictor of the response to anti-EGFR-TKI, a recent meta-analysis showed that wild-type EGFR patients can benefit from the therapy with an improved OS compared with placebo or standard chemotherapy [hazard ratio –0.930)] . TherefoBlood-based biomarkers in NSCLC are of particular interest as they can be easily and non-invasively accessed . Whole-gThe aim of the current study was to analyze blood-level exon array profiling data from unselected patients with advanced non-squamous NSCLC before and 24h after initiation of the combined targeted therapy BE. The specific objectives are twofold: (1) uncover which genes from whole blood circulating RNAs are immediately impacted by the effect of the combined therapy BE, and (2) assess the predictive value of these dysregulations.The gene expression data set originated from a translational substudy of the phase II SAKK 19/05 trial from the Swiss Group for Clinical Cancer Research . In the original study, 103 unselected patients with advanced non-squamous NSCLC were enrolled, among which 101 were evaluable. The experimental design of the trial SAKK 19/05 is summarized in Fig. RNA from whole blood samples was extracted and quality checked. Six pairs of samples had to be excluded from the analysis due to low quality, whereas RNA extracts provided sufficient quality for microarray hybridization in 43 out of 49 pairs of sample. Messenger RNAs were hybridized on Affymetrix Human Exon 1.0 ST arrays  following standard recommendations from the manufacturer. This microarray platform measures genome-wide exon-level expression in over 1.4 million probe sets, and allows the investigation of genomic variations both at the gene and at the exon level. For the sake of this analysis, 439,778 exonic probe sets  were kept in the analysis, after filtering out intronic, intergenic and unreliable probe sets (according to the nomenclature defined in the R package annmap ). Raw daThe current experimental design includes a repeated measurement of genome-wide exon-level expressions in 43 patients at two different time points (baseline and 24h after initiation of therapy). The exon expression data are stored in a pair of fully matched tables. Dually constrained correspondence analysis (DCCA)—an extension of the multivariate technique correspondence analysis—was used to investigate the 24h-change in exon expression levels . DCCA usGeneralized linear (mixed effects) models were used in order to test the predictive value of the identified biomarkers. Binary endpoints were tested using logistic regression, time to event data were modeled using Cox proportional hazards regression, and continuous variables were modeled using (multiple) linear regression. Mixed effects modeling was used when testing associations in the frame of the within-patient repeated measurement design. The adjustment for patients’ EGFR mutational status was done by including the mutational status as covariate in the predictive models. The significance level used for the discovery of the putative predictive markers was set to 0.001. Gene signatures combining the information of the best predictive candidates were built using the metagene approach . In thisHomo sapiens database version 86). Functions available in ade4  can theoretically be used to carry out DCCA. However, for reasons of computational efficiency due to the extensive size of exon array datasets, the DCCA algorithm was substantially optimized and a new R extension package dcca is available   and WikiThe characteristics of the 43 patients are reported in Table The 100 genes mostly dysregulated (54 up-regulated vs. 46 down-regulated) by the 24h effect of BE are summarized in Table  Based on the 100 best candidates identified by DCCA, three main pathways  were significantly altered by the 24h effect of the combined therapy BE: Hematopoietic cell lineage  and bevacizumab (“Hematopoietic cell lineage”). “Pathways in cancer” is a generic pathway, including genes involved in various aspects of tumorigenesis such as phenomena of proliferation, invasion, resistance and apoptosis. Several diseases are inter-related to this pathway, including non-small cell lung cancer. Most genes from the “Pathways in cancer”  are inhibited due to the action of BE resulting in an anti-proliferative and pro-apoptotic action of the combined therapy, with the notable exception of the up-regulation of the apoptosis inhibitor BIRC3. The combined action of BE results in the inhibition of all genes that belong to the “Hematopoietic cell lineage” . Hematopoietic stem cells play important roles for angiogenesis . The dowGene set enrichment analysis based on the WikiPathways database provides information on additional cancer- and energy-related activated pathways including “gastric cancer network 2” , RAD51 recombinase (RAD51), the junction plakoglobin (JUP), and the microphthalmia-associated transcription factor (MITF). E2F1 plays a critical role in the control of cell cycle and acts as a tumor suppressor. E2F1 was found to be associated with phenomena of resistance of targeted therapy in breast cancer . There wAll putative predictive markers of TTPBE at 24h were combined into a 91-gene metagene. Patients could be significantly classified into low-risk versus high-risk according to their median metagene score  were of small magnitude, yet consistent among patients. The use of a within-patient design of experiment including 2 time points before and after treatment helped to characterize these gene variations despite the relatively small sample size.The choice of the multivariate method DCCA over more common gene-by-gene approaches was driven by the fact that DCCA addresses the problem of the identification of differentially expressed genes (in within-patient repeated measures designs) in a multivariate manner. This is more statisfactory since it allows to take into account potential gene correlations/interactions using a single computationally efficient procedure. DCCA is an exploratory method appropriate for the purpose of the current hypothesis-generating translational study. On the other hand, gene-by-gene approaches are simple and flexible and could be preferred in case of more complex designs or when applied to confirmatory analyses.Although the magnitude of change of the most dysregulated genes over 24h was not predictive of the patient’s outcome, both the gene expression level at baseline and 24h revealed a series of putative predictive genes. While DS12 was defined as primary clinical endpoint of the original SAKK 19/05 trial, endpoints reflecting the activity of the treatment on the disease were more specifically investigated in the current translational substudy. TTPBE and TS12 are two endpoints which are objectively associated with the direct effect of BE. In both cases, a series of key predictive markers at 24h were enriched within the KEGG pathway “Pathways in cancer”. This pathway appears to play an important role both in the immediate effect of BE as measured in the blood, and in the prediction of the response to BE.Our findings could be validated using two independent datasets . The combination of the key predictive markers at 24h regarding TTPBE into a metagene was used to generate a gene signature, predicting with high significance patients into high vs. low risk populations. This signature was successfully validated, and could be used independently from the patient’s EGFR mutational status for proper patient selection.Because our gene signature is independent from the patient’s mutational status, it can be used as predictive marker both in EGFR mutated and wild-type populations. BE has potential to become a standard therapy in NSCLC patients with EGFR mutations, and our signature may help to select patients which may not respond to the therapy despite the presence of the mutation. Inversely, our signature may be useful for proper selection of BE responders among patients not harboring EGFR activating mutation.Our findings based on exon array data are in essence exploratory and future prospective confirmatory studies are needed to further validate the clinical relevance of our discovery.The 24h effect of BE could be accurately monitored in peripheral blood using the exon array technology. Genes impacted by the immediate effect of BE belonged to key signaling pathways, according to the expected mechanisms of action of both bevacizumab and erlotinib. Although the magnitude of change over 24h had no predictive value with regard to the investigated endpoints, the blood gene expression level measured 24h after initiation of BE could be used to predict TTPBE independently from the patient’s mutational status. Proper selection of responders to the combined targeted therapy BE could be monitored from blood level gene expression.


Patients with schizophrenia suffer from profound impairments of working memory and selective attention. These cognitive domains show a considerable overlap on both the behavioral and neurophysiological level. Importantly, selective attention appears to be crucial for the selection of information to be encoded into working memory. A number of studies have demonstrated that the efficiency of this “gatekeeper” function influences working memory performance. Furthermore, behavioural evidence indicates, that patients with schizophrenia have a specific deficit when required to suppress irrelevant but highly salient visual information during working memory encoding. Therefore, elucidating the neurophysiological mechanisms underlying the “gatekeeper” function of selective attention for working memory is highly relevant for understanding this deficit in schizophrenia. The aim of the current study was to investigate the neurophysiological correlates of encoding either salient or non-salient information in the presence of distractors of opposite saliency using functional magnetic resonance imaging (fMRI). Furthermore, we wanted to study the impact of additional top-down information guiding the selection of task relevant information.35 healthy volunteers underwent fMRI in a 3 T Siemens Trio scanner. During a change detection task four Gabor patches (two flickering and two non-flickering) with varying orientations were shown and participants had to memorise the orientations of the Gabor patches. A colored fixation cross was displayed before the stimuli either cueing two (predictive cue) or four (non-predictive cue) Gabor patch locations resulting in a 2 x 2 design of four conditions with the factors salience (flickering vs. non-flickering) and cue (predictive cue vs. non-predictive cue). During retrieval a single Gabor patch was displayed, and participants reported if the orientation was the same or had changed in that location. At the beginning of each block participants were instructed to either encode the flickering or non-flickering patches (targets) whose location could either be cued or uncued. In 80 % of trials, a target was probed during retrieval. Data analysis in Brain Voyager included standard data preprocessing. Additionally, a multiscale curvature driven cortex based alignment procedure was used to minimise macro-anatomical variability between subjects. Subsequently, functional data were analysed using a random-effects multi-subject general linear model . Functional connectivity analysis was performed using Granger Causality Mapping.Participants were able to preferentially encode task-relevant information in all four conditions. During encoding, they showed activation in a distributed network of fronto-parietal and visual areas. For salient compared to non-salient distractors, we observed increased functional connectivity between attention-related areas and extrastriate visual cortex. This difference was more pronounced for trials with a predictive compared to non-predictive cue.We were able to map the cerebral networks responsible for determining the contents of working memory. The observed patterns of connectivity indicate that core regions of the frontal-parietal network involved in both working memory and selective attention play a crucial role in the filtering of information by modulating the processing of information in visual areas. Our current findings provide the basis for studying the neurophysiological underpinnings of the interaction between impairments of working memory and selective attention in schizophrenia.


Short telomere length is a biomarker of cell oxidation and aging. Patients with first-episode psychosis (FEP) have been reported to have shorter telomeres than healthy controls (HC), suggesting that there is a premature and accelerated cellular aging in FEP. However, there are not data on longitudinal changes of telomere length in people with FEP relative to HC. We present preliminary results on 1-year longitudinal changes in peripheral blood mononuclear cells (PBMCs) telomere length and the proportion of PBMCs with short telomeres in young people with FEP and HC.16 young patients with FEP  and 21 young HC  were enrolled in the study. PBMCs telomere length and the proportion of PBMCs with short telomeres (i.e. <3kb) were determined using high-throughput quantitative fluorescence in situ hybridization (HT Q-FISH) at baseline (16 patients with FEP and 21 HC) and 12-month follow-up (4 patients with FEP and 4 HC).At baseline, we did not find significant differences in telomere length nor in proportion of PBMCs with short telomeres between FEP patients and HC. During the one-year follow-up, we found a significantly greater loss of telomere length  and a non-significantly trend for greater increase in the proportion of PBMCs with short telomeres  in patients with FEP than in HC.Telomere length changes during the first years of the illness can represent an early marker of accelerated cellular aging in patients with first-episode psychosis.


Eucalyptus nitens with only partially tracked pedigree. The sib-ship reconstruction allowed the identification of selfs (4% of the sample) and the exploration of their potential effect on inbreeding depression in the traits studied. We detected signs of inbreeding depression in diameter at breast height and growth strain while no indications were observed in wood density, wood stiffness and tangential air-dry shrinkage. After the application of a corrected sib-ship relationship matrix, additive genetic variance and heritability were observed to increase where signs of inbreeding depression were initially detected. Conversely, the same genetic parameters for traits that appeared to be free of inbreeding depression decreased in size. It therefore appeared that greater genetic variance may be due, at least in part, to contributions from inbreeding in these studied populations rather than a removal of inbreeding as is traditionally thought.Accurate inference of relatedness between individuals in breeding population contributes to the precision of genetic parameter estimates, effectiveness of inbreeding management and the amount of genetic progress delivered from breeding programs. Pedigree reconstruction has been proven to be an efficient tool to correct pedigree errors and recover hidden relatedness in open pollinated progeny tests but the method can be limited by the lack of parental genotypes and the high proportion of alien pollen from outside the breeding population. Our study investigates the efficiency of sib-ship reconstruction in an advanced breeding population of  Eucalyptus, where degrees of relatedness can vary widely due to mixed mating system ) in winter of 2014. Wood quality was measured on two different log lengths—log 1 from 1.4-3m and log 2 from 3-6m during winter of 2015. Wood traits assessed include density (WD [kg/m3]), wood stiffness  and growth strain  measured as log split width. Average tangential air-dry shrinkage was assessed only for log 2 (TS [%]). Wood density was measured as basic wood density through the maximum moisture content method . Wood stThe sib-ship inference was obtained by implementing the algorithm described in Wang (2004). This algorithm assumes markers to be selectively neutral, unlinked between loci and in linkage equilibrium. Additionally, they should have two or more codominant alleles and meet the assumption of Mendelian segregation. The sib-ship reconstruction was performed following the above described algorithm implemented in the COLONY package  using 50y is vector of measurements, β is vector of fixed effects representing intercept and seed orchard effects, a is the vector of random breeding values following A is the average numerator relationship matrix [r is the vector of random replication effect following I is the identity matrix and r(s) is the vector of random set nested within replication effects following e is the vector of random residuals following X and Z are the incidence matrices assigning fixed and random effects to measurements in y.The efficiency of relatedness recovery through sib-ship reconstruction on accuracy of genetic parameters such as additive genetic variance, heritability and genetic correlation was investigated with respect to their standard errors by implementing a linear mixed model within the ASReml-R statistical package  as follop matrix  developed is the vector of random dominance effects following D is the dominance relationship matrix created by setting the relationship coefficient of 0.25 between individuals belonging to the same full-sib family, diagonal elements were set to 1 for outbred individuals and 0.5 for selfs [Since the sib-ship reconstruction produced full-sib relationships see  section,Y is matrix of phenotypic measurements, a is vector of random additive genetic effects following var(a)∼N, where G1 is variance-covariance structure of additive genetic effects following st and 2nd trait, st and 2nd trait, r is vector of random replication effects following var(r)∼N, where G2 is variance-covariance structure for replication effects following st and 2nd trait, r(s) is vector of random effect of set nested within replication following var(r(s))∼N, where G3 is variance-covariance structure of set nested within replication effects following st and 2nd trait, e is a vector of random residual effects following var(e)∼N, where R is a variance-covariance structure for residual effects following st and 2nd trait, st and 2nd trait.A bivariate mixed linear model was used to estimate genetic correlations and test its benefit on accuracy of breeding values as follows:Two heritability estimates were obtained. Narrow sense heritability estimates were obtained using the original pedigree information or information from sib-ship reconstruction, and was estimated as follows:A second estimate was modified to include the estimated dominance variance using the dominance relationship matrix, and based on the reconstructed sib-ship structure as follows:th trait and th trait. Accuracies of breeding values were estimated as follows:i is inbreeding coefficient of ith individual [A is relationship matrix inferred from either the documented pedigree or the sib-ship reconstruction, c is vector of contribution where contributions were set equally for each individual as 1/n.Genetic correlations were estimated as the Pearson-product moment correlation as follows:dividual . The effdividual . Effectidividual  as folloE. nitens population. From within these, 500 random SNPs were selected for sib-ship analysis. The sib-ship reconstruction was performed in the COLONY package [The EUChip60k  provided package  and reco package  was estiWe compared the phenotypic performance of selfs and outbred individuals. Since there is the assumption that the selfing reduces genetic variance , the staEstimated heritabilities were low to moderate, ranging from 0.08 (DBH) to 0.45 (WD) and from 0.09 (DBH) to 0.34 (WD) in pedigree and sib-ship reconstruction based analyses. The heritability after sib-ship reconstruction decreased in most of the traits with the largest change from 0.41 to 0.28, observed in TS. By contrast, the heritability of DBH and GS2 increased after sib-ship reconstruction from 0.08 to 0.09 and from 0.18 to 0.22, respectively . Across When a dominance relationship matrix was included in the model, dominance variance was uncovered in ST1, ST2, and TS but was only found statistically significant in the last trait . HoweverMultivariate analysis based on pedigree information resulted in only a slight improvement in the accuracy of breeding values compared with results from the univariate analysis  and mostThe implementation of information from sib-ship reconstruction in multivariate analysis resulted in mixed results for estimation of genetic correlations, reflecting trends in additive genetic variance. Traits that showed a decrease in additive genetic variances resulted in reduced pairwise genetic correlations and vice versa. The highest improvement in genetic correlation estimated using sib-ship information was reached for traits suffering from inbreeding depression (DBH and GS2) . SimilarThe effect of sib-ship reconstruction in breeding values ranking was investigated with Pearson product moment correlations as well as with comparison of estimated genetic gains. The correlation between breeding values reached from 0.74 to 0.85 and corresponded to the changes in estimated genetic gain. The highest impact of sib-ship reconstruction was observed in DBH where the response to selection increased for 50% compared to pedigree based analysis . This isThe size, amount and precision of genetic parameter estimates dictates the level of genetic progress in any breeding activity. Heritability, an essential genetic parameter, is heavily influenced by the principle resemblance between relatives  and reliPedigree reconstruction  has beenE. nitens breeding program, where only the maternal side of the pedigree was tracked across all generations. Sib-ship reconstruction recovered the main portion of the pedigree-based half-sib relationships and also identified pedigree errors. Nevertheless, the methodology can only recognize major classes of relatedness such as unrelated, half-sib and full-sib pairs [Our study investigates a third generation of breeding population within the New Zealand ib pairs , which iib pairs  found atib pairs , 48. Capib pairs  tested dib pairs  proposedOpen pollination lacks inbreeding control and our sib-ship reconstruction recovered 27 individuals (4%) derived from selfing events, which largely contributed to a decrease in effective population size from 196.46 to 74.46. Selection of such individuals for the next generation of the breeding population would result in an undesirable increase in inbreeding level when no control over the paternal contribution is performed. This could potentially result in a considerable decrease of effective population size, needed for the long-term sustainability in genetic progress. Additionally, higher inbreeding levels may cause inbreeding depression, which in some populations/traits causes inferior phenotypic expression –53.E. nitens [Eucalyptus species [We investigated the phenotypic performance of individuals derived from selfing compared with individuals derived from outcrossing . The ave. nitens  which fo species . However species  which maMost of the traits showed a reduction or no change in heritability estimates and no change in accuracy of breeding values when sib-ship was used, which corresponds with findings in Vidal et al. . Other sThe recovery of full-sib families allowed us to construct a dominance relationship matrix, which was implemented in the mixed model to investigate non-additive genetic variance. Our analysis identified dominance variance in ST1, ST2, and TS but it was significant only in the last trait. The implementation of non-additive relationship matrices, especially dominance, improved the accuracy of genetic parameters in previous studies –59. HoweMultivariate analyses can further improve the precision of genetic parameters through genetic correlations , 64. OurThe precision of genetic correlations and the accuracy of breeding value estimates showed large improvement when information from sib-ship reconstruction was implemented. However, many genetic correlation estimates decreased and thus remained non-significant. Generally, about 100 families should be tested to obtain reliable and significant genetic correlations . The genOverall, the correspondence between breeding values produced in pedigree based and sib-ship reconstruction based model were between 0.74 and 0.85  which isIn addition, the estimated response to selection rapidly changed with change in heritability between investigated strategies. For traits suffering from inbreeding depression such as DBH, GS1 and GS2, the estimated genetic gain based on the original pedigree was largely underestimated. This was especially the case for DBH where the genetic gain when using sib-ship reconstruction increased for 50% compared with the scenario based on the original pedigree . In contOur analyses showed benefits from sib-ship reconstruction by identification of selfs to investigate the possible detrimental effect of inbreeding depression found in three studied traits. This allowed us to understand different patterns in genetic parameter changes when information from sib-ship reconstruction was implemented. Despite only rough inference about relatedness in advanced breeding populations due to an inability to recover the low classes of relatedness, sib-ship reconstruction provides a useful tool to recover major categories of relatedness where parental genomic information is missing. The technique will enable more effective management of genetic diversity and provide more precise estimates of genetic parameters compared with the previously incomplete documented pedigree.We found the implementation of sib-ship reconstruction resulted in higher heritability and genetic correlation estimates and improved accuracy of breeding values in traits showing signs of inbreeding depression. In traits free of inbreeding depression, the impact of sib-ship reconstruction was a decrease in heritability and only a negligible improvement in breeding values accuracy. We found lower correspondence between breeding values estimated from pedigree and sib-ship reconstruction, which can potentially bias the predictive ability in a genomic evaluation and prior pedigree reconstruction is recommended. Sib-ship reconstruction will be very useful implementation tool for breeders to ensure sufficient genetic diversity is present through inbreeding identification and eliminate selfs from selection in open-pollinated programs.


Comprehensive two-dimensional gas chromatography DeepwaterHorizon disaster that released approximately 160 million gallons of crude oil from the Macondo well (MW). Samples that were clearly collected following this disaster exhibit statistically significant match We validate our methods across 34 mentclass2pt{minimWe provide a peak-cognizant informational framework for quantitative interpretation of The online version of this article (doi:10.1186/s13065-016-0211-y) contains supplementary material, which is available to authorized users. Comprehensive two-dimensional gas chromatography A key distinction of our technique against multi-variate statistical methods  is compoTarget-based analysis –14: FocuTarget-agnostic analysis –22: StatAdditional file Many separation technologies routinely filter out non-target analytes, thus eliminating possibility of understanding their connection to dominant target analytes in an environmental sample. More comprehensive data sets recording the joint contributions of target and non-target analytes may be enabled through comprehensive two-dimensional gas chromatography entclass1pt{minimaibutions . Thus, centclass1pt{minimaDeepwaterHorizon disaster. The ROI biomarker region spans over a hundred compounds across a relative scale of 1−14.53 between the lowest and highest summits , a feature representation that collectively captures Topography partitions, a threshold-based partitioning technique for discovering source-specific and regional characteristics, andCross-PTM analysis, mathematical technique for directly determining “match” between two A natural outcome of PTM-based analysis is the discovery of topographic clusters (closely eluting groups of target and non-target biomarkers), which are key to understanding the regional and source-specific fingerprint.Our motivation in this work is to achieve robust forensic distinction between closely related oil sources by utilizing rich peak information diversity inmentclass2pt{minimchniques –22 that Deepwater Horizon disaster;Fourteen injections clearly originating from the MW, source of the Three injections from non-Macondo well oil originating from three different sources in the Gulf of Mexico; and,Seventeen injections from diverse oil sources outside the Gulf of Mexico region.In particular, injections 1 and 2 correspond to independent injections of a pre-spill sample taken directly from the MW during normal operations before the disaster; injection 3 corresponds to a surface slick sample from the MW collected after the spill; injection 4 is a post-spill sample collected directly from the broken riser pipe on June 21, 2010 [Deepwater Horizon disaster, and injection 17 is collected from a natural oil seep in the Gulf of Mexico in 2006. The remaining injections correspond to distant sources unrelated to the Gulf of Mexico. For example, injections 18, 19 and 20 are independent consecutive injections of the National Institute of Standards and Technology (NIST) Standard Reference Material 1582 .Additional file 21, 2010 , 35; injThe samples were analyzed on a  GC GC chromatogram of the MW pre-spill sample listed as sample #1 in Table S1. We focus on the hopane-sterane biomarker topography as the region of interest (ROI) as these compounds are well-known for their recalcitrance to environmental degradation [We introduce the PTM representation of mentclass2pt{minimradation , 36.PTM is a scalable node-based representation computed over a pre-selected p denotes the peak summit value, and m and n respectively denote the first and second dimension retention time indices for the particular peak in the Mathematically, each peak collapses into a single PTM node that stores two attributes: (i) the magnitude at the peak summit, and (ii) peak location. We represent information at a PTM node th node will be zero. Therefore, the problem of comparing two We note that the PTM matrices for the MW pre-spill sample . This peak ratio metric is calculated at the granularity of individual PTM nodes and assessed against a pre-selected threshold to decide a match between the test and reference samples for a given compound. These individual match assessments are then conducted across peak profiles spanning the The topography is partitioned into “similar” and “dissimilar” peaks that meet or fall below the match threshold. The percentage of peaks in the “similar” topography generates the cross-PTM score. The two partitions are called similarity and dissimilarity partitions, where similarity indicates the partition of the test In Additional file Therefore, we define a user-selected metric mentclass2pt{minimSimilarity partition: Portions of Dissimilarity partition: Portions of We note that either partition not only includes the peak summits, but also the region under a peak. In the scenario where a node was inserted in the test PTM  the We denote the The cross-PTM score, denoted as Deepwater Horizon disaster, including two pre-spill samples, and twelve post-spill samples collected at diverse locations after the Deepwater Horizon disaster, e.g. the plume at the base of the MW, grass blades on the Louisiana coastline, and oil slicks collected kilometers away from the disaster site (details provided in Table S1). These samples were collected in areas well documented [Deepwater Horizon disaster compared to the background.PTMs derived from cumented , 25 to bWe evaluate the cross-PTM score as a function of the peak ratio threshold across a diverse selection of injection pairs. We examine the robustness of intra-class match between injections of same origin against inter-class distinction between injection pairings from different origins. Specifically, we compare the fourteen MW injections (injections 1−14 in Table S1) against each other and against other sources within and outside the Gulf of Mexico region. We also compare the strength of MW vs. MW match against three other Gulf of Mexico injections (injections 15−17 in Table S1): (i) Eugene Island, (ii) Southern Louisiana Crude (SLC) and (iii) a Gulf of Mexico natural seep. Three consecutive injections from a non-Gulf of Mexico NIST sample originating in the Monterey area are also analyzed as an ideal intra-class case study, independent of any co-provenance bias with the Gulf of Mexico samples.Figure To provide a neutral baseline for best-case performance, we compare three NIST injections (injections 19–21 in Table S1), all of which were taken from the same sample of non-Gulf of Mexico origin. The NIST injections were run consecutively under practically identical experimental conditions. We observe in Fig. The 14 MW injections exhibit a range of 105–131 detected peaks spanning target and non-target Specifically, at the choice of This choice of peak ratio, We observe from Table As indicated earlier the proposed methods in chemometrics such as PCA can be applied towards quantitative The primary aim of this work is to provide quantitative peak-level interpretation beyond target biomarkers, with the end goal of robust differentiation between petroleum sources that share regional commonalities, and therefore, have highly correlated This differentiation between the two interpretation methods can be easily seen in Table The intra-class match (MW vs. MW) is slightly higher using PCA than PTM but the inter-class differentiation  is significantly more robust using PTM over PCA. This is to be expected as PCA is biased towards the common regional fingerprint of the Gulf of Mexico locale, which constitutes the dominant component of data variance of Mathematically, we can perform PCA cross-comparison between these correlated courses based on the non-dominant components, but these are typically vulnerable to baseline noise and other uncertainties, and as such, not reliable for robust source differentiation. This is evident in Fig. In summary, PCA enables statistical distinction between two In addition to results based on this experimental field data, we also provide extensive perturbation analysis of the PTM method over numerical simulations that introduce random variability of peak locations over the GCmentclass2pt{minimWe observed that the PTM approach is relatively immune to variability even when introduced variability is greater than the bounds In sharp contrast, the perturbation analysis of PCA scores over the same set of simulated images exhibit much higher “false alarm” match between classes, i.e., non-MW vs. MW comparisons. For example, the natural seep field sample was indistinguishable statistically from the Macondo class regardless of perturbation limits. PCA also exhibits much lower sensitivity to perturbations in the peak locations, which is to be expected, as it is a purely statistical compound-agnostic technique that does not consider one peak at a time. We note that the contrast between PCA and PTM observed over simulations is consistent with that observed over the field data.The PTM method enables We also wish to iterate that the PTM method has been developed in this work with recalcitrant biomarkers in mind. It certainly has the potential to apply beyond the recalcitrant hopane-sterane biomarker region, for other crude oils and distillates, by measuring changes in peak heights of the same compounds across samples collected at different times and locations. Ideally, we believe we will be able to quantify weathering processes, subtract them from the signal, and continue to make highly quantitative comparisons. However, such investigations warrant their own detailed study and as such, are outside the scope of this paper.Deepwater Horizon disaster, as well as over extensive perturbation analysis using numerical simulations  PTM, a feature representation that collectively captures the


Lake ecosystems are deeply integrated into local and regional economies through recreation, tourism, and as sources of food and drinking water. Shifts in lake phytoplankton biomass, which are mediated by climate warming will alter these benefits with potential cascading effects on human well-being. The metabolic theory of ecology suggests that warming reduces lake phytoplankton biomass as basal metabolic costs increase, but this hypothesis has not been tested at the global scale. We use satellite-based estimates of lake surface temperature (LST) and lake surface chlorophyll-a concentration  in 188 of the world’s largest lakes from 2002-2016 to test for interannual associations between chl-a and LST. In contrast to predictions from metabolic ecology, we found that LST and chl-a were positively correlated in 46% of lakes (p < 0.05). The associations between LST and chl-a depended on lake trophic state; warming tended to increase chl-a in phytoplankton-rich lakes and decrease chl-a in phytoplankton-poor lakes. We attribute the opposing responses of chl-a to LST to the effects of temperature on trophic interactions, and the availability of resources to phytoplankton. These patterns provide insights into how climate warming alters lake ecosystems on which millions of people depend for their livelihoods. If the availability of rate-limiting resources remains constant as lake ecosystems warm, resources will become scarcer relative to their demand causing a metabolic deficit3. Metabolic deficits at higher temperatures would leave lakes capable of supporting less phytoplankton biomass3. This theory has been used to explain how lake phytoplankton sizes and abundances decrease with warming in freshwater mesocosms4.Metabolic ecology has gained prominence, in part, for its capacity to explain and predict macroecological patterns and the influences of climate warming on the Earth. Based on the fundamentals of metabolic ecology, warming increases phytoplankton’s demand for resources to support higher metabolic rates at higher temperature5 or exacerbate6 the warming-induced metabolic deficit. Thus, the cumulative effects of warming on phytoplankton biomass remain uncertain for most of the world’s global population of large lakes. Uncertainty in the simple directionality of responses of phytoplankton biomass to temperature prevents meaningful estimates of how climate warming will affect lake carbon cycling7, food webs8, and biodiversity10 at the global scale. Understanding which lake attributes are associated with the strongest positive and negative effects of temperature on phytoplankton biomass could guide future research and inform lake management.However, warming has the potential to affect trophic interactions, and the availability of resources for phytoplankton, which could alleviate11, lake surface area11, and endemic lake species12. Even if the 188 lakes are biased relative to the global population of lakes in terms of their surface area, they are representative in terms of their depth, elevation, latitude, temperature, and average phytoplankton biomass. Changes in phytoplankton biomass in some large lakes have been shown to have substantial effects on species, economies, and livelihoods13.To address this uncertainty, we compared interannual variability in surface chl-a to LST for 188 of the world’s largest lakes (listed in Supplementary Table 14. We obtained daily chl-a concentration data from 2002-2016 from the Moderate Resolution Imaging Spectroradiometer (MODIS)-Aqua mission as pre-processed by the National Aeronautics and Space Administration’s (NASA) Ocean Biology Processing Group (OBPG). We merged the chl-a data with daily LST data from 2002-2016 from the Group for High Resolution Sea Surface Temperature (GHRSST) . We calculated average lake-wide Kendall’s rank correlations between chl-a and LST after accounting for seasonal and spatial variability within lakes (see methods). The average lake-wide correlations reported here reflect interannual associations between chl-a and LST, not seasonal associations, and are thus most likely to reflect the directionality of long-term lake temperature forcing. In situ monitoring data from the North American Great Lakes were used to validate the chl-a and LST data (see methods).Satellite measurements of lake colour provide a useful, high-resolution  phytoplankton biomass proxy that enables links to LST at broad spatial scalesp < 0.05; Fig. 15.We found that the lake-wide average correlations between chl-a and LST were highly variable across lakes (i.e. both positive and negative correlations) and a high proportion of correlation coefficients were significant , of which 74% (86 lakes) were significant after correcting for multiple comparisons  best explained variability in lake-wide average correlations between chl-a and LST.in situ data collected by the EPA  and phytoplankton-rich lakes (chl-a > ~20 μg/L). In phytoplankton-rich lakes, lake warming strongly favours phytoplankton species such as cyanobacteria which are less efficiently consumed by grazers19. The resulting reductions in trophic transfer efficiency from phytoplankton to their consumers may lead to an accumulation of phytoplankton biomass in warm years in phytoplankton-rich lakes20. However, community shifts toward less edible phytoplankton species are weaker in phytoplankton-poor lakes because cyanobacteria abundance in phytoplankton-poor lakes depends more on nutrient availability than on temperature alone21.Our results are consistent with previous work which suggests that the effect of warming on the trophic transfer efficiency of phytoplankton biomass to higher trophic levels depends on lake trophic state22 on phytoplankton25. Warming is expected to increase consumption rates, which would strengthen “grazer release” and promote higher phytoplankton biomass25. In contrast, where piscivores are present, chl-a may be low because of high grazing rates by primary consumers25. Consequently, warming induced enhancement of consumption rates would have the opposite effect of further reducing phytoplankton biomass in phytoplankton-poor lakes27. Thus, variability in the effect of warming on top-down limitation of phytoplankton biomass may partially explain why the correlations between chl-a and LST depended on lake trophic state  are common in the phytoplankton-poor regions of the tropical oceans36. Whereas the interannual correlation between chl-a and SST is more positive in the polar, relatively phytoplankton-rich regions of the marine environment37. As we have demonstrated for the world’s large lakes, these are not paradoxical responses because the indirect effects of temperature on phytoplankton in different contexts explain why opposite responses to warming can occur within the same ecosystem type.Our results are corroborated by 38. The simple band algorithms used here to convert lake colour data to chlorophyll-a concentration have been updated and improved in recent years39, but have not been calibrated for individual lakes. The optical properties of inland waters can make it difficult to use simple band algorithms to distinguish between chlorophyll-a and other dissolved and particulate substances (e.g. coloured dissolved organic matter)41. The development and validation of algorithms for optically complex waters could be substantially improved through rigorous validation against in situ data across the full spectrum of inland water types. However, research groups currently have access to in situ data from only a limited range of lakes. Regardless, our validation against in situ data from the North American Great Lakes suggests that our results may be robust to deviations between satellite and in situ data  which could elicit spurious correlations. Thus, we urge caution in interpreting the correlation coefficient in any specific lake as a strict warming effect. However, these independent drivers of both temperature and chl-a are likely to be lake-specific, thus we doubt that they would consistently bias our results comparing across many globally distributed lakes.Another important caveat is that our results reflect patterns at the surfaces of lakes during ice-free and cloud-free periods only. The relationship between chl-a and LST may vary with depth, ice cover, and cloud cover in ways that diverge from patterns shown here. While we have considered phytoplankton biomass as a response to temperature throughout our manuscript, phytoplankton sizes and abundances can also influence water temperature through its effect on light penetration in the water columnOur results may apply broadly to the global population of lakes despite our results being based on a subset of only the largest lakes, because lake surface area did not have a strong influence on its correlation between chl-a and LST. The relative influence of surface area in the boosted regression trees was only 9% which is less than the null expectation given eight variables in the model (12.5%). However, according to the boosted regression trees, smaller lakes did have slightly more positive correlation coefficients between chl-a and LST than larger lakes. Extrapolating this trend to smaller lakes would indicate that chl-a and LST may be more positively correlated on average for the complete global population of lakes. Future work using lake colour data from satellites with finer spatial resolution (e.g. Sentinel-3A) should enable direct tests of how warming affects phytoplankton biomass in smaller lakes. Merged chl-a data products which combine measurements from multiple sensors may also enhance the spatial and temporal coverage for smaller lakes, enabling the inclusion of more lakes in future studies. Our computational approach also required more data per lake than other less computationally intensive approaches would have required. Simpler approaches using linear models instead of boosted regression trees could increase the number of lakes with sufficient data for model fitting.7, lake warming rates43, lake ecology45, and lake-derived benefits to humans46. Higher phytoplankton biomass in phytoplankton-rich lakes may exacerbate problems associated with anthropogenic lake nutrient enrichment, such as the expansion of anoxic zones, harmful algal blooms, fish die-offs, and reduced water clarity. Managers may need to reduce anthropogenic nutrient loads that were acceptable in the past to maintain ecosystem functions in phytoplankton-rich lakes as they warm48. In contrast, the reduction of phytoplankton biomass in phytoplankton-poor lakes with warming presents its own potential management challenges such as reduced fisheries productivity45. For instance, fish production in Lake Tanganyika has already been substantially diminished as a result of climate-mediated reductions in phytoplankton biomass and production6. In some cases, managers may want to fertilize lakes to sustain fish production45, but managers must prudently weigh the changing costs and ecological risks of those actions. Thus, the amplification of lake trophic states with warming may require adaptive lake management efforts at the local level to prevent loss of lake benefits to humans. Otherwise, human livelihoods are likely to be affected across gradients in lake trophic state at the global scale.Changes in phytoplankton biomass that result from lake warming are likely to affect lake carbon cycles49. GHRSST data at 1 km resolution are based upon night time skin and sub-skin surface temperature observations from the NASA Advanced Microwave Scanning Radiometer-EOS (AMSRE), the Moderate Resolution Imaging Spectroradiometer (MODIS) on the NASA Aqua and Terra platforms, the US Navy microwave WindSat radiometer, Advanced Very High Resolution Radiometer (AVHRR) on several National Oceanic and Atmospheric Administration (NOAA) satellites, and in situ SST observations from NOAA. Daily estimates of lake surface chl-a were obtained at 4 km resolution from the MODIS Aqua mission dataset processed to Level-3 data by the Ocean Biology Processing Group50 of NASA. Chl-a data are generated using the OCI band ratio algorithm based on the method of Hu et al.39. Chl-a and LST data covered the time period from July 2002 to November 2016.Daily estimates of lake surface temperature were obtained from the blended, level-4 data product of the Group for High Resolution Sea Surface Temperature (GHRSST) version 4.151. Satellite data were available from only a subset of these water bodies because many lakes were too small to be included or were persistently obstructed by clouds or ice.To ease the computational intensity of this work, we used 0.1-degree median-filtering followed by data subsetting to a spatial resolution of 0.1-degrees for all analyses. We also used a 5-day median filter followed by data subsetting to a temporal resolution of 5 days for all analyses. Lake sections were identified using the level 1 Global Lakes and Wetlands Database (GLWD-1) comprising the 3067 largest lakes and 654 largest reservoirs worldwide including basic attribute data rlake), we first calculated correlation coefficients separately for every latitude-longitude-day of the year combination in our dataset. To do this, we subdivided all observations in the 15-year time series by its pixel (resolution of 0.1 degrees). We further subdivided the data from each pixel by the day of the year on which the data were measured. As a result, each subdivision was composed of up to 15 observations of chl-a and LST—one observation for each of the 15 years in the time series. For subdivisions with at least eight coincident observations of chl-a and LST (following minimum sample size recommendations from published literature52), we used non-parametric Kendall’s rank correlation to determine the interannual relationship between chl-a and LST . We used non-parametric Kendall’s rank correlations to assess the relationship between LST and chl-a to avoid spurious correlation coefficients in cases where the relationship between in situ and satellite data are nonlinear or where the linear slope was not equal to one.To estimate the average lake-wide correlation coefficient between chl-a and LST (rlake) using the correlation coefficients calculated for every latitude-longitude-day of the year combination (ri). However, the ri’s were not randomly distributed over space and season. So, to avoid biasing rlake toward the season or location in space with the most ri’s, we used boosted regression trees to factor out those biases. To do this, we fit boosted regression trees separately for each lake which modelled the observed ri’s as a function of latitude, longitude, and day of the year with a total of n observations (ri’s). Boosted regression trees were used because they allow for nonlinear relationships between independent and dependent variables and high levels of interactions among independent variables (e.g. the effect of season could depend nonlinearly on latitude and longitude simultaneously). To reduce the computation intensity of this work, models were fit to a random subset of n = 10,000 observations for lakes which had more than 10,000 ri’s  from the distribution of residuals from the boosted regression trees without replacement. rlake was calculated as the mean rlake was significantly different from zero.We calculated each lake’s average lake-wide correlation coefficient between chl-a and LST  following definitions from published literature53. We had an incomplete predictor matrix with some missing data (see Supplementary Information), but boosted regression trees can accommodate partially missing data without dropping observations from the model. We weighted each rlake in the model by the significance of its p-value  so that less significant correlations had less influence on the model outcome. To visualize the effect of median chl-a alone on the correlation between chl-a and LST, we removed variability attributable to all other predictor variables  from the North American Great Lakes. In situ data used here were collected in the North American Great Lakes primarily during two annual field campaigns in early and late summer from 2002–2016. We merged the EPA’s in situ data with the nearest coincident satellite data and directly compared them using standard major axis regression (SMA). We found that for all lakes, in situ chl-a and LST were significantly correlated to satellite-based chl-a and LST . R2 from the SMA for the relationship between in situ and satellite chl-a and LST were 0.78 and 0.99, respectively. The median absolute error for chl-a and LST were 0.26 mg m-3 and 0.44 °C, respectively. The slopes in the SMAs for chl-a varied from 0.66 (Lake Erie) to 1.23 (Lake Ontario) across the North American Great Lakes due to differences in their optical conditions . The slopes in the SMAs comparing remotely-sensed to in situ LST varied from 0.98 (Lake Ontario) to 1.06 (Lake Superior) across the North American Great Lakes  with an SMA slope not significantly different from 1 . Lake-wide correlations between chl-a and LST were also highly correlated  with an SMA slope not significantly different from 1 . These results show that even if the relationship between in situ and remotely sensed chl-a is at times weak . In situ temperature and chl-a data analysed in the current study are available through the United States Environmental Protection Agency (EPA) Central Data Exchange (https://cdx.epa.gov/). Lake polygons and characteristics are available through the Global Lakes and Wetlands Database .All satellite-derived LST and chl-a datasets analysed during the current study are publicly available through NASA’s Physical Oceanography Distributed Active Archive Center (Supplementary Information


Carotid body tumors typically arise at the level of the common carotid bifurcation where the carotid body exists. Superior and inferior extension beyond the carotid body may occur as well, especially if the tumor is long-standing. We herein describe a case of carotid body tumor in a patient who presented with a right-sided neck mass for 30 years. Computed tomography angiography (CTA) and B-mode sonography with color-coded Doppler sonography showed a vascular tumor arising at the level of the right common carotid artery bifurcation with superior and inferior extension beyond the bifurcation. This report emphasizes the utility of computed tomography angiography (CTA) and B-mode sonography with color-coded Doppler sonography as non-invasive modalities in visualizing the extension of carotid body tumors. Paragangliomas are rare tumors that arise from widely dispersed specialized neural crest cells that are associated with autonomic ganglia. Carotid body tumors represent 60 - 70% of head and neck paragangliomas -2. A carWe describe herein a case of a 53-year-old male who presented with a painless neck mass that he first noticed 30 years ago. The initially small mass has progressively grown and is now accompanied by night sweats and dizziness. The patient denies blurry vision, parasthesia, paralysis, palpitations, difficulty swallowing, or changes in voice.Physical examination showed a right neck mass about 4 x 3 cm. It was non-tender, and does not move with swallowing. No lymphadenopathy was appreciated. Cranial nerves I-XII were grossly intact except for decreased sensation in his right lower mandible and the superior right aspect of his neck.CT angiography of the neck demonstrated an avidly enhancing mass splaying the right external carotid artery (ECA) and the internal carotid artery (ICA). The mass begins 1.6 cm below the right CCA bifurcation with the bulk of the tumor measuring 4.3 x 3.4 x 5.8 cm. There was additional tumor projecting approximately 1.6 cm above the dominant mass. The right internal jugular vein is compressed .B-mode sonography showed a solid, well-defined, and hypoechoic tumor splaying the ICA and ECA . Color fCarotid body tumors can be diagnosed by B-mode sonography with color-coded Doppler sonography, computed tomography (CT), magnetic resonance imaging (MRI) and digital substraction angiography (DSA). B-mode sonography typically shows a solid, well-defined, and hypoechoic tumor. Color Doppler imaging in most cases, but not all, reveals intratumoral hypervascularity. B-mode sonography and color Doppler imaging can both display the splaying of the carotid bifurcation with displacement of the external carotid anteriorly and both the internal carotid and the internal jugular vein posteriorly -4.CT Angiography is capable of showing the highly vascular tumor as a well-defined soft-tissue mass within the carotid space of the infrahyoid part of the neck with characteristic splaying of the carotid bifurcation. The homogeneous and intense enhancement following intravenous administration of contrast material is due to the underlying hypervascularity of the tumor. Splaying of the carotid artery bifurcation has classically been described on conventional angiography and is known as “Lyre” sign -7. This Digital Subtraction Angiography (DSA) is an invasive modality to diagnose carotid body tumors, but is more important for therapeutic purposes. It provides endovascular access for possible tumor embolization and identifies the blood supply and flow dynamics of the tumor. This would enable the surgeon to locate displaced vessels and identify more unpredictable and less accessible feeding vessels. Moreover, later phases of the arteriogram visualize the venous outflow, which provides information to the surgeons so that they can preserve the venous drainage until the final stages of tumor mobilization which results in decreased intraoperative blood loss .In summary, ultrasound is a non-expensive, non-invasive modality that can be used as a first step to diagnose carotid body tumors. CT Angiography and MRI can show the extension of the tumor. Conventional angiography is an invasive modality for diagnosis, but is more important for embolization and preoperative evaluation. Our case emphasizes the utility of computed tomography angiography (CTA) and B-mode sonography with color-coded Doppler sonography as non-invasive modalities in visualizing the extension of carotid body tumors.


Conditional transgene expression in human stem cells has been difficult to achieve due to the low efficiency of existing delivery methods, the strong silencing of the transgenes and the toxicity of the regulators. Most of the existing technologies are based on stem cells clones expressing appropriate levels of tTA or rtTA transactivators (based on the TetR-VP16 chimeras). In the present study, we aim the generation of Tet-On all-in-one lentiviral vectors (LVs) that tightly regulate transgene expression in human stem cells using the original TetR repressor. By using appropriate promoter combinations and shielding the LVs with the Is2 insulator, we have constructed the Lent-On-Plus Tet-On system that achieved efficient transgene regulation in human multipotent and pluripotent stem cells. The generation of inducible stem cell lines with the Lent-ON-Plus LVs did not require selection or cloning, and transgene regulation was maintained after long-term cultured and upon differentiation toward different lineages. To our knowledge, Lent-On-Plus is the first all-in-one vector system that tightly regulates transgene expression in bulk populations of human pluripotent stem cells and its progeny. VP-16) linked to the TetR repressor (tTA for Tet-Off or rtTA for Tet-On systems). The presence of this transactivating domain can make these proteins toxic due to the sequestration of transcription factors required for cell growth (squelching)1314pseudo-TetO sites can trans-activate non-target cellular genes161819loxP systems are useful tools for conditional transgene expression, they have the potential to influence cellular physiology.Technologies allowing conditional transgene expression in human stem cells are fundamental not only to study gene function145681011Another major obstacle for the wider application of most conditional systems is the general requirement of drug selection to generate drug-responsive clones that can regulate transgene expression. The generation of regulatable stem cells clones is not always possible  and, when possible, is time-consuming and labor-intensive. In this direction, efficient genetic manipulation of stem cells is a critical aspect to achieve direct transgene regulation. The gene delivery system must achieve stable expression of the regulator and long-term regulation of the transgene in target stem cells and in its progeny. The main hurdles to achieve this goal in stem cells are the low efficiency of gene delivery methods and the strong silencing of the transgenes2226However, although LVs are one of the most efficient systems to achieve stable transgene expression in stem cells, they are also prompt to transgene silencing2829303132343336Our group has previously described an all-in-one regulated lentiviral vector (CEST) based on the original TetR repressor, that allowed the generation of Dox-regulated cell lines, including primary human fibroblasts (HFF) and human MSCs (hMSCs) by repression of the strong CMV promoter. However the CEST LVs was unable to regulate transgene expression in pluripotent stem cells and required multiple integrations per cell in order to achieve regulation in 293 T and hMSCsIn order to generate Tet-On all-in-one LVs that tightly regulate transgene expression in human stem cells we have constructed several LVs harboring different modifications . All theNext, we constructed four all-in-one LVs harboring different promoters and insulator combinations ; The CESExpression of the TetRnl2 through the hEF1α promoter in 293 T cells decreased the leaking of the vectors in the absence of Dox 404243As expected, the CESTnl2Is2 LV showed improved expression levels compared to their Is2-negative counterpart (CESTnl2) in the presence of Dox (+ (−Dox)/%eGFP+(+Dox)] × MFI eGFP+ cells (−Dox)] to take into account the expression levels of the transduced cells in the absence of Dox  *100/%eGFP+(+Dox) that generate values from 0 (no leaking) to 100 (absence of regulation). However, this analysis does not discriminate between partially-responsive cells from those cells that do not respond to Dox. We have therefore reanalyzed the leaking using an alternative formula; leaking =  or [[% eGFP+ (−Dox) x 100/%eGFP+(+Dox)]] x MFI eGFP+(−Dox) (supplementary information). Background (%eGFP+ cells in MOCK transduced cells) were subtracted to the % eGFP+ obtained in the different transductions and conditions. Therefore when the %eGFP+ (−Dox) is identical in Mock and Transduced cells the leaking is zero and when the %eGFP+ (−Dox) is identical to the %eGFP+(+Dox) the leaking is 100.Transduced cells incubated in the presence or absence of Dox (0.1 μg/ml) were analyzed by flow cytometry to determine the percentage and Mean fluorescence intensity (MFI) of eGFP positive cells as well as the MFI of WLC (WLC = P1 defined as the 7-AAD negative population). The fold induction was calculated using the following formula MFI P1 (+Dox)/MFI P1 (−Dox). Similarly, the The promoter methylation was assessed by bisulfite treatment of Genomic DNA and sequencing of the resulting converted gDNA. In the presence of sodium bisulfite, only the unmethylated cytosines are chemically converted to uracil. The genomic DNA was isolated from hESCs at the desired time points using the DNaseasy kit . Sodium bisulfite treatment of genomic DNA was performed using the EpiTect bisulfit kit (Qiagene), according to the manufacurer’s instruction. The converted gDNA was used for a nested PCR amplification of SFFV and EF1a promoters. The primers used for each amplification is shown in Suplementary Information . The PCRwww.graphpad.com) applying the unpaired two-tailed t-test. Statistical significance was defined as a P value < 0.05.All data are represented as mean +/− (SEM). The statistical analysis was performed using the GraphPad Prism software .Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


Autophagy is a tightly regulated lysosomal degradation process that mediates the sequestration of intracellular entities to form autophagosomes, and their delivery to lysosomes for bulk degradation .Autophagy plays a key role in the maintenance of cellular homeostasis, in baseline conditions as well as in response to stress. It can function as either a pro-death or pro-survival mechanism in cancer cells, depending on the different stage of tumor development and the cell type . During Cellular energetics represents a critical regulatory point for autophagy. Indeed, 5’ adenosine monophosphate-activated protein kinase (AMPK) senses the equilibrium between the phosphorylation status of the adenine nucleotide pool. When equilibrium is shifted to low phosphorylation of nucleotides , AMPK is activated and promotes autophagy to recycle energetic constituents. Mitochondria are a major site of production of higher energy state adenine nucleotide . The hig2+ transfer between ER and mitochondria supports mitochondrial respiration and then suppresses autophagy through low AMPK activity [This signaling platform participates in the regulation of autophagy. Caactivity .Additionally, the ER-mitochondria interface contributes to autophagosome formation, and proteins localized at MAM compartments are indispensable for proper autophagic vesicle formation .Several oncogenes and oncosuppressors localize at MAMs to regulate communication between organelles and alter cell proliferation or sensitivity to cell death stimuli .2+-dependent apoptosis by blocking the activity of IP3R3 and promoting the formation of a multiprotein complex containing IP3R3, AKT and the protein phosphatase PP2a [Taken together, these observations led us to hypothesize about the possible involvement of promyelocytic leukemia protein (PML) in autophagic process . PML is ase PP2a . In our Pml-/- mouse embryonic fibroblasts (MEFs) and in the livers and skeletal muscles of adult Pml-/- mice compared with WT conditions; moreover, loss of PML increased autophagosome biogenesis.We observed increased abundance of LC3-II in Pml-/- MEFs. In contrast, a control construct targeted to nuclei failed. Given the tight interaction between PML and p53 and that p53 localizes at ER/MAMs [In particular, PML localization at MAMs is necessary for repressing autophagy. In fact, an erPML construct (which contains the entire PML protein targeted to the outer surface of the ER) suppressed the elevated levels of autophagy when re-introduced into  ER/MAMs , we sougWhat are the molecular mechanisms involved in PML-dependent autophagy regulation? We examined the relationship between PML and a multiprotein complex that includes AMP-activated protein kinase (AMPK), mammalian target of rapamycin (mTOR) and unc-51-like kinase 1 (Ulk1) that is emerging as an important player in the regulation of autophagy.2+ transfer is important for induction of autophagy given the new critical role of Ca2+ in the formation of autophagosomes.Moreover, we speculate that downregulated ER-mitochondria Ca2+ from the ER into the mitochondria and the production of ATP are reduced [In the absence of PML, the release of Ca reduced .Pml-/- cells and tissues exhibited increased AMPK phosphorylation levels, Ulk1 hyper-phosphorylation and reduced phosphorylation of mTOR and its substrate p70S6K, leading to increased autophagy. Interestingly, restoration of Ca2+ signaling in a Pml-/- background by overexpressing mitochondrial Ca2+ uniporter (MCU) restores basal levels of autophagy.Our results demonstrated that These data indicate that the loss of PML from MAMs confers an alternative pathway to sustain mitochondrial activity and thus cell survival during stress conditions, promoting cell survival in the tumor environment.in vivo models. We observed that the absence of PML promotes tumor development associated with resistance to anticancer drugs due to increased autophagy levels in the tumor. Pharmacological inhibition of autophagy through chloroquine administration restored chemotherapy-related apoptosis (Figure We further investigated the cross-talk between autophagy and PML-related cell death in s Figure .Our findings hold therapeutic implications for the treatment of solid tumors associated with PML down-regulation and allow for further consideration for the MAM microenvironment that could be a fundamental background for the regulation of autophagic-dependent cancers.