Bevacizumab (Avastin) has rapidly gained status as a broadly active agent for malignancies of several different histologies in adults. This activity has spawned a range of uses in pediatrics for both oncologic and non-oncologic indications. Early analyses indicate that pediatric cancers exhibit a spectrum of responses to bevacizumab that suggest its activity may be more limited than in adult oncology. Most exciting, is that for low-grade tumors that threaten vision and hearing, there is not only evidence for objective tumor response but for recovery of lost function as well. In addition to oncological indications, there is a range of uses for non-oncologic disease for which bevacizumab has clear activity. Finally, a number of mechanisms have been identified as contributing to bevacizumab resistance in cancer. Elucidating these mechanisms will guide the development of future clinical trials of bevacizumab in pediatric oncology. Angiogenesis, or the formation of new blood vessels from pre-existing vessels is essential for normal organogenesis and tumorigenesis. The wide range of requirements for new blood vessel growth during development and in the setting of pathology is served by numerous angiogenic factors that recruit endothelial progenitors and other pro-angiogenic and vascular supporting cells, and promote endothelial cell proliferation and survival. Vascular endothelial growth factor (VEGF) is essential to both normal and pathology-associated angiogenesis and consequently, VEGF antagonism is the primary focus of anti-angiogenic therapies. Multiple isoforms of VEGF are generated through alternate splicing and post-translational modifications of the products of a multi-gene family comprised of VEGF-A, -B, -C, -D and placenta growth factor Ferrara, . Most of121, VEGF165, VEGF189, VEGF206)  as well as two co-receptor, Neurophilin 1 and 2. While binding to VEGFR-1 occurs with higher affinity than binding to VEGFR-2, it is VEGFR-2 that mediates the pro-angiogenic effects of VEGF on endothelial cells  in patients who had progressed after prior treatment  every 2 weeks. Fifteen of the patients had no objective sustained response. Eight of the patients with GBM had stable disease for ≥12 weeks with a median of 176 days and median PFS of 4.2 months. Median PFS for patients with DIPG was 2.3 months. This study concluded that there is a lack of efficacy of combined bevacizumab and irinotecan in children with recurrent GBM and DIPG.Outcomes for children with high-grade gliomas (HGG) remain dismal with 5-year-survival rates of 5–15% for grade IV gliomas and 20–40% for grade III gliomas  every 2 weeks. Seven had a radiographic response  and seven had clinical improvement . At the conclusion of the study, eight of the nine patients had not progressed . The 14th patient was progression-free at 45 months. Significant responses to bevacizumab in children with LGG is consistent with reported activity for other anti-angiogenic therapies such as thalidomide-based regimens  are the most common childhood brain tumor. Treatment varies as a function of age and tumor location between surgical resection alone, or combined surgery and chemo or radiation therapy. Common indications for adjuvant therapy include recurrence in surgically inaccessible locations and threats to function particularly for deep-seated midline tumors growth of which can result in vision loss, or hypothalamic or brainstem dysfunction. While LGG are slow growing, biologically benign tumors, they possess increased vasculature . Although benign, these tumors tend to inexorably progress and often result in loss of all functional hearing by middle age. Standard treatment includes surgery and radiation however, these approaches are frequently ineffective at preserving hearing . Importantly, four of the seven evaluable patients had 57% improvement in word recognition beginning approximately 8 weeks after initiation of therapy and four of those five also reported improvement in tinnitus. Mautner et al.  treated with bevacizumab (5 mg/kg every 2 weeks for 12 months)  had a significant reduction in both MRI fluid-attenuated inversion-recovery (FLAIR) and T1-post-Gd contrast abnormalities. The average daily dexamethasone dose was reduced by 8.6 mg. A retrospective review at Children’s Hospital of Denver evaluated the response to bevacizumab of patients with DIPG (n = 4) , a severe complication of allogeneic stem cell transplantation. Yabe et al.  reportedVascular endothelial growth factor is a potent inducer of vascular permeability. Blocking it in patients with severe edema such as radiation necrosis and CLS may benefit some patients. Further studies are needed to further evaluate these findings.Bevacizumab has been used in several case reports of patients with angiomatosis, a neoplasm-like entity. Smith et al.  reportedBevacizumab has also been reported to benefit a patient with multifocal lymphangiomatosis or Gorham–Stout disease (GSD), presenting as idiopathic osteolysis of the bones. GSD is possibly triggered by occult neoplastic cells that produce large amounts of IL-6 and VEGF. Grunewald et al.  reportedp = 0.002). Treatment of ROP with bevacizumab is now the standard of care at many centers. Albini and Murry reported a retrospective, non-comparative, open-label, interventional case series in 35 eyes of 33 patients with diseases such as CVN, Coat’s disease, familial exudative vitreoretinopathy and sickle cell retinopathy who were treated with bevacizumab . One patient with sickle cell retinopathy and central retinal vein occlusion had significant improvement in central macular thickness over 6 months after one injection (811–56 μm). Similar effectiveness was also reported for treatment of CNV after Stevens–Johnson syndrome , choroidal neovascularization (CNV), Coat’s disease, and familial exudative vitreoretinopathy can lead to severe vision loss. Vision loss can occur for many reasons, including retinal ischemia and VEGF-induced neovascularization and vascular leakage of fluid, blood, and exudates. Mintz-Hittner et al.  conducteMaturo and Hartnick  reportedn = 1) including anaphylaxis, cerebral ischemia, posterior reversible encephalopathic syndrome, proteinuria, lymphopenia, and pneumonia.In adult studies, serious adverse events have been reported with the use of bevacizumab including CNS hemorrhage (1–5%), hypertensive crisis (<1%), and blood clots (6–9%) (Scappaticci et al., In the cases reported by Hwang et al.  who wereIn patients with NF2 treated with bevacizumab at 5 mg/kg/dose, no thromboembolic events, hemorrhage, congestive heart failure, gastrointestinal perforation, reversible posterior leukoencephalopathy, or Grade III/IV events have been reported (Plotkin et al., Secondary amenorrhea was reported in a few patients. In the Phase I COG study, two of three post-menarchal females had a rise is LH and FSH, one subsequently missed her periods (Glade Bender et al., The patient treated for CAT syndrome was noted to have multiple punched out boney lesions after four doses of bevacizumab (Smith et al., Together, these studies indicate that bevacizumab is generally well tolerated in the pediatric population at doses up to 15 mg/kg/dose. Serious side effects seen in adults have not been reported in the pediatric population.Targeting VEGF pathways clearly has a role in the treatment of a broad range of human disease. Some applications, like the treatment of post-irradiation edema or CLS, relate more to VEGF-mediated regulation of vascular permeability rather than to VEGF-stimulated angiogenesis. In these circumstances bevacizumab has demonstrated significant effect and it is likely that its use as a single agent for these indications will increase. In contrast, as detailed above, in cancer treatment when the angiogenic functions are the critical target for VEGF directed therapies, bevacizumab fails in most cases to produce a lasting clinical response. Identifying the mechanisms of resistance and appropriately modifying treatment approaches will be essential for the full realization of the potential of anti-angiogenic therapies. Here we will briefly review proposed mechanisms by which cancers adapt and evade VEGF antagonism.In discussion of and investigation into mechanisms of bevacizumab resistance, much has been made of an apparent change in the pattern of recurrence in bevacizumab-treated, versus conventionally treated, GBM. Some retrospective and prospective studies have reported an increase in the frequency of diffuse and distant recurrence after bevacizumab therapy from approximately 10–50% (Norden et al., The mechanisms that drive resistance to bevacizumab therapy are not yet fully defined. However, preclinical and clinical studies have revealed multiple intratumoral and systemic pathways that can contribute to tumor progression on bevacizumab therapy and these have been well reviewed by Bergers and Hanahan . In respIn addition to developing therapeutics that can target the multiple mechanisms of angiogenesis and tumor recurrence, a great clinical challenge as we advance anti-angiogenic therapies for cancer will be the application of laboratory and imaging techniques that can better define response early in therapy. This will be especially critical in the application of anti-angiogenic therapies to brain tumors where serial biopsy is not currently available. In this regard, serum evaluation for the up-regulation of pro-angiogenic factors, especially those associated with resistance like VEGF, PIGF, PDGF, FGF, IL8, CXCL12 may provide insight into shifts in response (Murukesh et al., In addition, imaging, particularly MRI-based spectroscopy, perfusion, diffusion, and oxygen consumption sequences may be able to detect functional changes in the vasculature relevant to anti-angiogenic responses and recurrent angiogenesis (Pope et al., Finally, PET imaging for therapy-induced changes in expression of specific components of angiogenic pathways as well as for changes in vascular function is an additional promising approach for detecting response to anti-angiogenic therapies that is just entering clinical evaluation (Chen and Chen, Together, sophisticated and combinatorial monitoring of response to anti-angiogenic therapies may support the timing of adjunctive interventions to circumvent resistance mechanisms and enhance efficacy.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The MUS-81, SLX-1, and XPF-1 structure-selective endonucleases have been implicated in meiotic crossover (CO) formation in a variety of organisms, but their contributions to C. elegans CO formation have been unclear. In this issue of PLOS Genetics, Agostinho et al., Saito et al., and O'Neil et al. demonstrate that MUS-81 and XPF-1 function in two parallel pathways during the formation of meiotic crossovers in C. elegans and provide important insights into the interplay between endonucleases and the Bloom helicase ortholog during crossover formation.Saccharomyces cerevisiae have provided extensive genetic and biochemical support for this model. Although initiation by DSBs appears to be universal  and a progression of joint molecule (JM) intermediates that link DNA duplexes on homologous chromosomes  and S. cerevisiae Sgs1. Sgs1 disassembles early meiotic JMs, both to generate noncrossovers and to prevent formation of aberrant multi-chromatid JMs et al. report that him-6 mutants have defects in processing early intermediates, leading to chromosome fragmentation et al. propose that HIM-6 also has a late function in generating COs in conjunction with XPF-1 Drosophila BLM Additional potential similarities arise from studies of C. elegans meiosis  might work together to resolve HJs. However, it is possible that the proteins act on a different structure, such as nicked HJs (e.g., see reference Another important question concerns the exact nature of the pre-crossover JM. The authors suggest that these resolvases are acting on HJs, and resolution of chromatin bridges by GEN1 supports this suggestion. Agostinho


Kenneth Hugdahl's second affiliation is Department of Radiology, Haukeland University Hospital, Bergen, Norway.lower degrees of language lateralization as determined with fMRI (van Ettinger-Veenstra et al., higher degrees of lateralization.”On page 1 the final sentence in the second column should read: “Moreover, individuals with n = 16 to find correlations between ear asymmetry and behavioral language tests in the non-forced condition of the Bergen DL task.”On page 7 the final sentence of the first column should read: “For the same reason van Ettinger-Veenstra et al.  might habut see Catani et al., On page 8 the final paragraph of the discussion should read: “As far as language is concerned, however, stronger lateralization seems to be associated with better performance in verbal abilities (Boles et al.,


Was−/− mouse. Both cellular and humoral immune defects in WAS patients contribute to the onset of severe clinical manifestations, in particular microthrombocytopenia, eczema, recurrent infections, and a high susceptibility to develop autoimmunity and malignancies. Autoimmune diseases affect from 22 to 72% of WAS patients and the most common manifestation is autoimmune hemolytic anemia, followed by vasculitis, arthritis, neutropenia, inflammatory bowel disease, and IgA nephropathy. Many groups have widely explored immune cell functionality in WAS partially explaining how cellular defects may lead to pathology. However, the mechanisms underlying the occurrence of autoimmune manifestations have not been clearly described yet. In the present review, we report the most recent progresses in the study of immune cell function in WAS that have started to unveil the mechanisms contributing to autoimmune complications in WAS patients.Wiskott–Aldrich Syndrome (WAS) is a severe X-linked Primary Immunodeficiency that affects 1–10 out of 1 million male individuals. WAS is caused by mutations in the WAS Protein (WASP) expressing gene that leads to the absent or reduced expression of the protein. WASP is a cytoplasmic protein that regulates the formation of actin filaments in hematopoietic cells. WASP deficiency causes many immune cell defects both in humans and in the WAS murine model, the  WAS gene is located on the X chromosome and encodes a 502 amino acid protein  is also an important regulator of WASP activation by inducing a stable acting form  is a rare X-linked Primary Immunodeficiency (PID) that affects 1–10 out of a million male individuals  is observed in some patients with substantial protein expression  cells  in T cells and T Cell Receptor (TCR)-dependent activation  and fungal infections  as compared to WASP-positive patients , followed by vasculitis , arthritis (29%), neutropenia (25%), inflammatory bowel disease (9%), and IgA nephropathy (3%). Henoch–Schönlein-like purpura, dermatomyositis, recurrent angioedema, and uveitis have also been reported in some patients , most available therapies are not curative. Intravenous immunoglobulins (IVIG) and antibiotic prophylaxis are often used to reduce the risk of infections in WAS patients, but it is not clear whether these treatments effectively reduce the incidence of life-threatening infections  is available, BMT leads to 73–100% survival . This adverse event gives rise to some concerns on the safety of RV-mediated GT for WAS. A multicenter clinical trial using a third generation LV carrying WAS gene driven by the endogenous promoter is on going in Milan, Paris, and London. Preclinical data in the murine model indicate that the LV-mediated GT approach is effective in restoring immune cell functionality . Two different Gene Therapy (GT) clinical trials have been approved: a Retroviral Vector (RV)-mediated gene transfer  family member Fas ligand (FasL) that is released and binds its receptor Fas in an autocrine fashion , Multiple Sclerosis (MS), and Systemic Lupus Erythematosus . In the last decade, it has been clearly defined that the lack of WASP causes defects in the cytoskeletal functions of B cells, including adhesion, migration, and homing  and a reduced or absent antibody production to polysaccharides and other T cell-independent antigens  co-engagement in Was−/− B cells from chimeras could mediate tolerance breakdown, since the loss of Myeloid Differentiation primary response gene 88 (MyD88) signaling abolished the production of anti-dsDNA antibodies, germinal center formation, and development of systemic autoimmune disease , Recher et al.  which in turn activate pDCs to produce high levels of type I IFN , in the pathogenesis of systemic autoimmune diseases. In particular, IFN-α produced by pDCs upon recognition of foreign nucleic acids via TLR7 and TLR9 contributes to tolerance breakdown in several autoimmune diseases, such as SLE, SS, and psoriasis (Ronnblom, Was−/− mice abolish the production of anti-dsDNA antibodies (Becker-Herman et al., −/low B cells (Ng et al., − B cells in WAS patients (Park et al., Was−/− mice (Westerberg et al., Triggering of autoreactive B cells by self-nucleic acid-containing ICs can be another possible mechanism underlying the production of autoantibodies in WAS. In fact, it is known that self-nucleic acid-containing ICs can activate B cells through synergistic engagement of BCR and TLR7 or TLR9 (Leadbetter et al., In conclusion, together with the defects already described in the literature, these future lines of enquiry underline the greater than expected extent to which the WASP deficiency affects the immune system. Further research is necessary to define the underlying molecular and cellular mechanisms leading to autoimmunity, which represents the main collateral damage caused by WASP deficiency.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.


While Frye and Jackson do a good job describing the common mechanisms of resistance found in food animals in the U.S., they err in stating that in the U.S. extended spectrum β-lactamases (ESBLs) “thus far have only been found in human and not food animal isolates.”Escherichia coli expressing CTX-M-type ESBLs in fecal isolates from healthy cattle at a livestock market and from a diagnostic isolate submitted to the Ohio Animal Disease Diagnostic Laboratory. Since then, CTX-M-type ESBLs have been found in E. coli collected from healthy dairy calves in the western U.S. (Davis et al., Salmonella enterica isolates from swine in Minnesota and from turkeys in 4 states (Wittum et al., E. coli from swine finishing barns in Michigan and Ohio and in Klebsiella pneumoniae from swine in Illinois (Mollenkopf et al., In fact, Wittum et al., in 2010 first reported the collection of E. coli isolate expressing CTX-M-type ESBLs from Pennsylvania has been reported (Doi et al., E. coli expressing OXA-type ESBLs (Davis et al., We are not aware of studies finding CTX-M-type ESBLs in isolates from broiler chickens, but a retail chicken meat


MTHFR gene includes a common polymorphism (rs1801133 or C677T), which is associated with enzyme activity. The T-allele of the C677T polymorphism has been associated with earlier age at onset of schizophrenia in a Scandinavian population, although no association was found in replication attempts in other populations. Extending the study to five Nordic samples consisting of 2,198 patients with schizophrenia, including the original Scandinavian samples, there was no significant association between MTHFR C677T polymorphism and age at onset in schizophrenia. The present results do not suggest that the investigated MTHFR polymorphism has any significant influence on age at onset of schizophrenia in the Nordic population. © 2012 Wiley Periodicals, Inc.Methylenetetrahydrofolate reductase (MTHFR) is an enzyme involved in metabolic pathways of importance for nucleotide synthesis and methylation of DNA, membranes, proteins and lipids. The  Methylenetetrahydrofolate reductase (MTHFR) is an enzyme involved in metabolic pathways of importance for nucleotide synthesis and methylation of DNA, membranes, proteins and lipids [Frankenburg, MTHFR gene includes two common polymorphisms  which both alter enzyme activity and homocysteine concentrations [Frosst et al., MTHFR C677T among 40 polymorphisms associated with schizophrenia susceptibility in the Han Chinese population [Yue et al., The MTHFR C677T functional polymorphism, and the metabolism of folate, methionine, and homocysteine have been extensively studied in relation to etiologically complex chronic diseases, and a relationship between MTHFR C677T and age at onset have been observed in for example, coronary artery disease, breast cancer, and Parkinson disease [Mager et al., MTHFR 677 T-allele predispose to an earlier age of onset in unrelated patients with schizophrenia of Scandinavian origin and found similar results in a family-based sample from western China [Vares et al., MTHFR polymorphisms in unrelated patients with schizophrenia to a combined analysis on age at onset. However, we were unable to replicate the findings outside the initial Scandinavian sample [Vares et al., The MTHFR C677T polymorphism and age at onset in two additional samples of Icelandic and Danish patients with schizophrenia, and combine the results with the previous Scandinavian results in a meta analysis.To determine whether the initial finding in unrelated individuals was restricted to populations of Scandinavian origin, or more likely due to chance alone, we here analyse the relationship between The Icelandic sample consisted of patients with schizophrenia, who were recruited all over Iceland as previously described [Stefansson et al., The sample from Denmark/Aarhus has also been previously described [Demontis et al., The samples from the initial report [Vares et al., P < 1 × 10−5, or frequency difference between chip types or typing centers with P < 1 × 10−6 were also excluded. Only samples with yield >90% were included, and the lower yield of each pair of duplicates was removed. Markers had control HW equilibrium P values > 0.001, and, in each group, yield in cases and controls was >95%.Genotyping of the Icelandic group using Illumina Human Hap300Beadchip genome-wide arrays was carried out as previously described [Stefansson et al., In Denmark/Aarhus, DNA was extracted from dried blood spots provided by the Danish Newborn Screening Biobank using the Extract-N-Amp Blood PCR kit . DNA was subsequently whole-genome-amplified using the RepliG mini kit  and genotyped on the Illumina Infinium HD Human610-Quad BeadChip  as previously described [Hollegaard et al., MTHFR C677T SNP was genotyped at the SNP Technology Platform in Uppsala, Sweden (http://www.genotyping.se), using the Illumina BeadStation 500GX and the 1536-plex Illumina Golden Gate assay (Illumina Inc.) as previously described [Vares et al., In the initial Scandinavian sample [Vares et al., For each study, we estimated the allele association between the C677T polymorphism and age at onset of schizophrenia with a general linear model. In this primary analysis, age at onset was treated as a quantitative trait, and modeled as a function of gender and the number of T alleles . The analyses were conducted in parallel, with R for the Icelandic and Danish/Aarhus sample  and with SAS/STAT® software (version 9.3) for the previously investigated Scandinavian samples [Vares et al., Meta analysis of the five studies was done with a simple random-effects model with Proc Mixed in the SAS/STAT® software . In the analysis, the allele association was modeled as a function of the fixed effect of the intercept and the random effect of study. The beta coefficients (regression slopes) for C677T from the primary analyses were used as the observed effect size, and the corresponding squared standard error (SE) was treated as known variance in the diagonal of the variance–covariance matrix R. HW equilibrium was tested using Fisher's exact test as implemented in PEDSTATS [Wigginton and Abecasis, MTHFR 677T allele frequency varied from 27% (Sweden) to 36% (Iceland) in the samples . A negative association between 677T and earlier age at onset was also apparent in the Norwegian sample, although in this sample the association strength was weaker . We noted that this sample consisted of patients with an earlier age of onset than the other Nordic samples, and most patients became ill before the effect of the MTHFR C677T polymorphism was apparent in the original Scandinavian samples [Vares et al., MTHFR C677T polymorphism and age at onset of schizophrenia in the Icelandic sample (P = 0.86). Thus the initial finding of an age dependent association between the MTHFR C677T polymorphism and age at onset of schizophrenia in the Scandinavian population could not be replicated in two large and independent samples of similar origin. A meta-analysis of all five samples resulted in a pooled association estimate of 0.42 ± 0.26 years earlier age at onset of schizophrenia per T-allele (mean ± SE), which did not reach statistical significance (P = 0.17).In the independent Danish sample (Aarhus), the age at onset decreased with on average 0.25 years per MTHFR 677T-allele and age at onset of schizophrenia in two independent samples of northern European heritage (Denmark and Iceland), or in meta-analysis including the initial Scandinavian samples. These results are in line with earlier analyses outside Scandinavia [Peerbooms et al., In the present study, we were unable to replicate our original finding of a negative association between the MTHFR 677T-allele and earlier age at onset in our original Scandinavian sample was primarily due to the Danish (Copenhagen) sample, and a similar tendency was present in the Danish (Aarhus) replication sample. Age at onset varied among the cohorts included in the study, and part of this variation may have been related to the different definitions of age at onset used . The heterogeneity in the definitions of age at onset is likely to have increased phenotype variation (and thus decreased the power to detect a true but weak association), but we observed no relationship between the definition of age at onset and association strength, neither in this nor in our previous analysis of multiple samples [Saetre et al., MTHFR polymorphism with respect to varying environmental conditions, such as availability of folic acid in food and/or practices of folic acid supplementation during pregnancy. Moreover, the power to detect an effect of the MTHFR 677T allele on late onset schizophrenia would be limited in the Danish (Aarhus) sample simply because of its younger age structure. Thus we cannot exclude the possibility of a weak negative association in populations of North European origin.The association between MTHFR 677T-allele has no significant effect on the age at onset of schizophrenia in the general population, irrespective of the geographical location, ethnicity or genetic background. Nevertheless, given the important role of MTHFR in the metabolism on folic acid, methionine, and homocysteine, it is possible that MTHFR gene variation not only affects schizophrenia susceptibility but may also modify symptom severity [Herran et al., However, given the data at hand, and previous analysis of samples of different geographical and ethnical origins [Peerbooms et al.,


Phosphodiesterase-5 (PDE5) inhibitors have shown a beneficial effect in a variety of clinical conditions, such as benign prostate hyperplasia, pulmonary arterial hypertension, female sexual arousal disorder, overactive bladder, and incontinence, Raynaud's disease, heart failure and stroke among others  pools (see Figure in vivo models of prostate cancer (Das et al., The accumulating body of evidence suggests that PDE5 inhibitors could interfere with the efflux functions of the ABC transporters, thus sensitizing cancer cells toward cytotoxic agents that are substrates of ABC transporters (Ding et al., In summary, there is evidence that suggests that PDE5 inhibitors may have an anticancer action either by increasing cGMP-PKG and coupled downstream events or by their ability to inhibit ABC transporter—mediated s efflux of anticancer drugs. The safety, high tolerability, and wide availability of PDE5 inhibitors have made this class of drug an attractive tool to investigate their role in cancer chemotherapy. Since a detailed understanding of PDE5 inhibitors as anticancer adjuvants is limited, further studies are warranted to characterize their mechanisms and establish their role.


DNA methylation is one of the essential epigenetic mechanisms that are closely correlated with the mechanisms underlying cell growth, differentiation and transformation in eukaryotes. Global changes in the epigenetic landscape are considered to be a hallmark of cancer. The initiation and progression of cancer are mediated through epigenetic modifications along with genetic alterations. Aberrant methylation of promoter regions is an epigenetic abnormality of the human genome that is highly characteristic of cancer. In this review, we aimed to summarize our current understanding of the alterations in the epigenetic landscape and investigate the potential use of DNA and RNA methylation in effective molecular treatment strategies. Complex diseases, such as malignant tumors and diabetes, are a common occurrence and represent a major public health concern. Despite the significant advances in cancer treatment, the overall cancer-related mortality is ~90%, due to late-stage diagnosis and failure to optimize treatment. Therefore, effective biomarkers for cancer diagnosis are urgently needed.Cancers such as lung, colon and breast cancer are frequently diagnosed at a late stage. Despite intensified efforts focused on improving the survival of cancer patients, only a moderate improvement is generally achieved. Failure of early diagnosis often leads to low treatment efficiency and poor prognosis; thus, the identification of a signal characterizing the early stages of formation and progression of cancer may reduce the incidence of this disease . Early fThe mechanisms underlying cancer development are complicated and cancer was originally perceived as a genetic disease. However, it was previously demonstrated that the initiation and progression of cancer involve epigenetic abnormalities, such as DNA methylation, histone modifications, nucleosome positioning and aberrant expression of non-coding RNAs, specifically microRNAs –7. The tcis-acting elements with trans-acting factors and gene expression. Various epigenetic experiments, involving genomic imprinting . Chromosome remodeling or reactivation of transposable elements was generated via demethylation of exons and introns or repetitive DNA sequences. An increased number of DNA hypomethylation loci in CpGs was shown to increase chromosomal instability and oncogene activation. For example, the hypomethylation of LINE-1 and Alu retrotransposons is frequently associated with lung cancer . The distribution of m5C within DNA is unique and may be used for genome-scale methylation analysis . For exaAs a specific pattern of gene expression in mammals, DNA methylation is essential for tissue development. Abnormal DNA methylation commonly disrupts molecular signaling mechanisms and leads to the development of various diseases, such as cancer. DNA methylation was the first epigenetic alteration to be identified in cancer . Therefoet al  were found to harbour methylated sites (ESR1 and 14-3-3-σ promoters were significantly different compared to healthy controls (et al (et al (The DNA methylation status is correlated with cancer and the methylation level is inversely correlated with mRNA expression levels. Jin et al  reporteded sites . In breacontrols . Glocknes (et al  reportedl (et al  comparedCDH1, APC, MGMT, RASSF1A, GSTP1, p16 and RAR-β2, affect the activity of tumor suppressor genes, typically leading to transcriptional silencing. A number of important genes that undergo silencing interfere with important cancer-related cell pathways. Thus, the aberrant methylation of the promoters of cancer-related genes may be deemed as a potential biomarker contributing to early cancer detection and prediction of prognosis.The identification of patients with organ-confined carcinoma is key to early-stage diagnosis of cancer. Common organ-confined cancers, such as lung, hepatic, breast, cervical, colorectal and genitourinary tract cancers, result in patient death due to the lack of effective clinical diagnosis. In these cancers, the methylation of the promoter regions of tumor suppressor genes, such as SEPT9, is commonly detected in the plasma of patients with primary CRC and was submitted to the FDA for marketing application in 2010 as a molecular marker for early clinical stage CRC. SEPT9 as a DNA methylation biomarker was also associated with breast cancer; Gonzalez et al (SEPT9 expression may contribute to the pathogenesis of certain types of breast cancer. Another potential biomarker for breast cancer is the methylation of PITX2. The evaluation of the PITX2 methylation status among different breast cancer patient populations successfully increased the outcome prediction performance. Harbeck et al (PITX2 methylation in 399 breast cancer specimens and identified low-risk patients with hormone receptor-positive and node-negative disease. Hartmann et al (PITX2 in 241 breast cancer specimens and concluded that methylation of PITX2 was correlated with clinical outcome. Hrasovec et al (TMEM25 were correlated with CRC, with TMEM25 hypermethylation possibly playing a significant role in altering the expression of this gene in CRC.A series of novel DNA methylation biomarkers in the plasma and stool were developed for various detection purposes. Vimentin is transcriptionally silent in normal epithelia and aberrant vimentin expression has been used as a cancer marker in fecal DNA testing . In 2005ez et al  indicateck et al  investignn et al  also anaec et al  reportedThe elucidation of the mechanisms that underlie DNA methylation changes in cancer cells may help identify a number of cancer-specific methylation markers, assisted by promising detection methods, ultimately resulting in optimized clinical applications. The realization that DNA methylation may be involved in human malignancies and is ubiquitous in human diseases is likely to promote the development of novel diagnostic, preventive and therapeutic strategies.et al (et al (et al (RASSF1A to be an epigenetic marker for the detection of fetal DNA in maternal plasma. Epigenetic information passed from parent to offspring and DNA methylation from gametes was shown to be predominantly maternal (et al (The association between methylation and heredity is currently an emerging research topic. The presence of modified m5C affecting the phenotype of the offspring may be parentally inherited. Carone et al  indicatel (et al  reportedl (et al  considermaternal . Similarl (et al  recentlyThe modified DNA m5C that extensively occurs within the genome was extensively investigated over the last few decades; however, numerous putative methylated RNAs have been identified and characterized, regarding location, mechanism of formation and cellular function . Additio


Almost 40 years ago, Takatsuki et al. recognized the existence of a peculiar T cell leukemia in Kyoto, Japan that they named Adult T Leukemia (ATL). They reported a series of 13 patients in 1976 (Uchiyama et al., Almost 10 years ago, ours colleagues Kuan Teh Jeang and Mitsuaki Yoshida organized a special issue on HTLV infection in Oncogene. In setting up this issue, we cannot forget the memory our friend Teh.We called upon the expertise of different research groups from Europe, Japan, and USA. However, we regret that the format of this issue prevented us from soliciting many other colleagues. The following reviews will deal with many fascinating aspects of viral cycle, but summarizes also new approaches that should allow a better integrated research.A first group of articles provides information about HTLV-1 epidemiology and associated-pathogenesis. The article from Gessain and Cassar provides an updated view on HTLV-1 distribution, based on data obtained from 1.5 billion individuals originating from endemic areas (Gessain and Cassar, A second group of articles summarizes the interaction between the virus and the host's cells. Before causing diseases, HTLV-1 has to enter the cell. However, the mechanisms of HTLV-1 transmission and cell entry have remained elusive for a long period of time. Pique and Jones have summarized recent insights about those mechanisms both at the cell level but also between individuals (Pique and Jones, in vivo (Bai and Nicot, A third group of articles reports data on individual viral proteins that play important roles in the viral cycle and/or in pathogenesis. Nakano and Watanabe remind us the important role played by Rex, which uses cellular pathways to export unspliced or singly spliced viral mRNAs in the cell cytoplasm, therefore allowing expressing of structural proteins (Nakano and Watanabe, Finally, Duc Dodon and colleagues remind us that studying HTLV-1 pathogenesis requires animal models (Dodon et al.,


Frontiers in Neural Circuits in response to the “Special topic” entitled “Cortical NO interneurons: from embryogenesis to functions.” Although it is practically impossible to address all aspects of NO interneurons, this special issue organized in four sections covers different facets of neocortical and hippocampal NO interneurons: from their diversity and embryonic origins to their functions in the cortical circuits and in neurovascular coupling.Neuronal processing and physiology of cortical circuits rely on a delicate interplay between glutamatergic excitatory neurons and GABAergic inhibitory interneurons in a spatially, temporally, and cell-type specific manner. Understanding these processes is further complicated by the large diversity characterizing the cerebral cortex (Ascoli et al., Studying the functional roles of NO-producing cortical interneurons has been challenging. NO being a highly diffusible gas (Wood and Garthwaite, stratum radiatum of the CA1 area exhibit characteristic properties of ivy cells and express NPY. In contrast to their counterparts in the stratum pyramidale (Fuentealba et al., stratum radiatum ivy cells display a different pattern of axonal and dendritic arborizations suggesting that these two subpopulations of hippocampal NO interneurons are involved in different microcircuits.This issue was revaluated in the whisker-to-barrel cortex by using NADPH-diaphorase histochemistry (Nogueira-Campos et al., Concomitantly with improvements in nNOS detection and the advent of genetic tools allowing lineage analysis of interneuron precursors, the developmental temporal and spatial origins of hippocampal and cortical nNOS neurons have been clarified with three studies showing that hippocampal and cortical type I NO interneurons originate from the medial ganglionic eminence (Jaglin et al., Another important question addressed in this special topic, is how NO interneurons are modulated by afferent signals and what their main neuronal targets are. Type I NO interneurons were found to express selectively the NK1 receptor whose activation by its natural agonist substance P induced their depolarization (Dittrich et al., Although NO is a well-established vasodilator, the relevance of NO interneurons in neurovascular coupling, the tight relationship between neuronal activity and local blood perfusion, has been questioned for decades (Cauli and Hamel,


Brain-derived neurotrophic factor (BDNF) is a potent regulator of neuronal development and synaptic plasticity that is fundamental to neural circuit formation and cognition. It is also involved in the control of appetite and body weight, with mutations in the genes for BDNF and its receptor, TrkB, resulting in remarkable hyperphagia and severe obesity in humans and mice. Recent studies have made significant progress in elucidating the source, action sites, and regulatory pathways of BDNF with regard to its role in the control of energy homeostasis, and have shed light on the relationships between BDNF and other molecules involved in the control of body weight. Here we provide a comprehensive review of evidence from pharmacological, genetic, and mechanistic studies, linking BDNF to the control of body weight. This review also aims to organize the main findings on this subject into a more refined framework and to discuss the future research directions necessary to advance the field. NTR). The Trk receptors have different neurotrophins as their preferred ligands, with NGF binding to TrkA, BDNF and NT4/5 binding to TrkB and NT3 binding to TrkC, while all of the neurotrophins can bind to p75NTR  is a member of the neurotrophin family of secreted signaling molecules that includes nerve growth factor (NGF), neurotrophin-3 (NT3), and neurotrophin-4/5 (NT4/5). Signal transduction can occur through the binding of two distinct classes of receptor proteins: the tropomyosin receptor kinase (Trk) family of receptor tyrosine kinases, which includes TrkA, TrkB, and TrkC, or the p75 neurotrophin receptor , dorsomedial hypothalamus (DMH), ventromedial hypothalamus (VMH), paraventricular hypothalamus (PVH), and brainstem Flier, . The ARCBdnf genes have nine exons, the first eight of which contain 5′ untranslated regions (5′ UTRs), with exon IX containing the entire coding sequence as well as the 3′ untranslated region (3′ UTR) , ventral tegmental area (VTA), substantia nigra and nucleus of the solitary tract (NTS) , after which the signal peptide is cleaved, leaving proBDNF  mice, which had increased risk of obesity resulting from increased food intake, concomitant with elevated serum leptin and insulin levels   infusion of neurotrophins on presynaptic cholinergic function  is the major site of thermogenesis. Sympathetic inputs from the stellate ganglion increase the expression of uncoupling protein 1 (UCP1) in IBAT, which allows the energy generated from β oxidation of fatty acids to dissipate as heat in response to physiological stimuli such as cold exposure and overeating  expressing Cre recombinase indeed caused modest hyperphagia and obesity  was used to delete the Bdnf gene in SF1-expressing VMH neurons, the resulting mutant mice did not display any body weight phenotype (Dhillon et al., Cre transgenic mice will greatly facilitate elucidation of the role of VMH BDNF in the control of feeding behavior and body weight.The VMH is likely a site for the synthesis of BDNF protein that is critical for energy homeostasis. The Bdnf gene expression.Food deprivation also decreased levels of BDNF protein in the rat DVC, whereas refeeding or peripheral injection of the anorexigenic hormones leptin or CCK increased BDNF protein levels in this area (Bariohay et al., TrkB gene expression was reduced to a quarter of the normal amount, or the Bdnf gene was deleted in CaMKIIα-expressing neurons, displayed markedly increased food intake on high fat diet compared to low fat diet (Xu et al., Bdnf mRNAs in the VTA and deletion of the Bdnf gene in the VTA induced hyperphagia on high fat diet but not low fat diet (Cordeira et al., Mice in which Bdnf mRNA encoding the same protein due to the presence of two alternative polyadenylation sites: short 3′ UTR Bdnf mRNA and long 3′ UTR Bdnf mRNA. One study indicates that BDNF protein regulating food intake is translated from long 3′ UTR Bdnf mRNA, likely in dendrites (Liao et al., Bdnf polyadenylation site, such that the long Bdnf 3′ UTR is truncated (Gorski et al., Bdnf mRNA (An et al., Bdnf mRNA in the VMH and nearby regions completely prevented these animals from developing hyperphagia and obesity (Liao et al., Bdnf mRNA to regulate appetite.Neurons produce two forms of NTR (Reichardt, NTR, lead to obesity in humans and mice (Lee et al., Not much is known about the action sites of BDNF with regard to its role in the control of energy homeostasis. BDNF exerts its biological effects through two receptors, TrkB and p75TrkB mouse knockouts. Once the neuronal populations are identified, researchers can focus on the mechanisms by which BDNF modulates the function of these neurons.Pharmacological studies have suggested several putative sites where BDNF acts to control energy homeostasis. Direct injection of BDNF into either the PVH or the VMH of adult rats was found to decrease food intake and increase energy expenditure (Wang et al., TrkB mRNA in the VTA (Cordeira et al., 1, such that peripheral injection of a D1 receptor agonist normalized the hyperphagia on high-fat diet (Cordeira et al., 1 is likely indirect, as the vast majority of TrkB-expressing neurons are striatal medium-sized spiny neurons that express the dopamine receptor D2 (Baydyuk et al., In addition to the hypothalamus and the brainstem, the mesolimbic dopamine system may be another target for BDNF to affect food intake, especially hedonic eating. Consumption of high fat diet was found to increase It is worth noting that one study reported that while central administration of TrkB agonists was anorexigenic in non-human primates, similar to what has been observed for rodents, peripheral administration was orexigenic, opposite to what is observed for rodents (Lin et al., db/db mice, diet-induced obesity mice and lethal yellow agouti mice, peripheral administration of BDNF has been shown to prevent or ameliorate the diabetic and obese phenotypes (Nakagawa et al., db/db mice, BDNF could potentiate the action of insulin in peripheral tissues, reducing serum glucose in obese, diabetic db/db mice (Ono et al., db/db mice by affecting blood glucose control, rather than food intake. BDNF administration also normalized fasted blood glucose levels, possibly through reduced hepatomegaly (Tonra et al., db/db mice, suggesting a long-lasting effect of BDNF on the control of glucose metabolism (Ono et al., db/db mice were restored to normal levels by BDNF treatment, with increased islet beta-cell area in BDNF-treated mice and a reversal of the decrease in pancreatic secretory granules, indicating a role for peripheral BDNF in restoring impaired pancreatic insulin production and secretion in db/db mice (Nakagawa et al., Obesity in mice with BDNF deficiency is associated with hyperglycemia and impaired glucose tolerance (Kernie et al., db/db mice led to increased insulin receptor activation in the liver and insulin-stimulated PI3K activity in the liver, skeletal muscle and IBAT. However, no direct effect of BDNF was found on cultured hepatocytes, L6 muscle cells or 3T3-L1 adipocytes, suggesting an indirect route of action. This may possibly be through regulation of central mechanisms leading to peripheral signaling, as suggested by the observation that centrally administered BDNF also had hypoglycemic effects, and led to similar increases in liver insulin receptor activation and insulin-stimulated PI3K activity (Nakagawa et al., db/db mice, indicating the activation of sympathetic pathways to regulate glucose metabolism (Yamanaka et al., CNS neurons may mediate some effects of peripheral BDNF administration on glucose homeostasis. Chronic subcutaneous BDNF administration in Bdnf mRNA levels in the VMH (Komori et al., Bdnf mRNA in dendrites, as leptin increased dendritic Bdnf mRNA translation in cultured hypothalamic neurons (Liao et al., Bdnf mRNA and BDNF protein, likely by activating BDNF-expressing neurons through a polysynaptic neural circuit.Several studies indicate that BDNF acts downstream of anorexigenic factors such as leptin and CCK to regulate food intake and body weight. Peripheral leptin administration was found to increase Bdnf mRNA (Liao et al., Bdnf mRNA, leptin injection activated LepRb normally in the ARC, DMH, and VMH, as indicated by phosphorylation of STAT3; however, leptin-induced c-Fos expression was impaired in the ARC and VMH and abolished in the DMH (Liao et al., BDNF signaling through the TrkB receptor is necessary for the anorexigenic action of leptin. Peripheral leptin injections reduced food intake in wild-type mice, but failed to do so in mice lacking long 3′ UTR TrkB mutant mice and Mc4r−/− knockout mice (Xu et al., TrkBF616A mice in which the phenylalanine residue at position 616 of the TrkB receptor is changed to an alanine residue, which makes the TrkB kinase sensitive to inhibition by the small molecule 1NaPP1 (Chen et al., Bdnf mRNA levels in the VMH (Xu et al., BDNF also interacts with the melanocortin system to regulate energy homeostasis. The initial consideration of this interaction arose from the appreciation of similar obesity phenotypes between Bdnf gene is a common allele (Shimizu et al., Cre transgenic mice and circuitry-mapping tools, it is becoming feasible to identify the neural circuits mediating the effects of BDNF on energy homeostasis. Identification of these circuits and elucidation of the mechanisms by which BDNF acts on them will greatly enhance our understanding of human obesity and hopefully bring us closer to a strategy for designing effective drug treatments for this disease.Strong evidence of a crucial role for BDNF in the control of energy homeostasis has been accumulated from genetic and pharmacological studies during the last decade or so. The Val66Met polymorphism in the The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The mechanisms of eNOS-uncoupling seem multiple and complex. Recent research provides emerging evidence supporting an essential role of increased activity of arginases including arginase-I and arginase-II in causing eNOS-uncoupling, which results in vascular oxidative stress and inflammatory responses, and ultimately leading to vascular diseases. This review article will summarize the most recent findings on the functional roles of arginases in vascular diseases and/or dysfunctions and the underlying mechanisms in relation to oxidative stress and inflammations. Moreover, regulatory mechanisms of arginases in the vasculature are reviewed and the future perspectives of targeting arginases as therapeutic options in vascular diseases are discussed.Oxidative stress and inflammation in the vascular wall are essential mechanisms of atherosclerosis and vascular dysfunctions associated with risk factors such as metabolic diseases, aging, hypertension, etc. Evidence has been provided that activation of the vascular endothelial cells in the presence of the risk factors promotes oxidative stress and vascular inflammatory responses, leading to acceleration of atherosclerotic vascular disease. Increasing number of studies from recent years demonstrates that uncoupling of endothelial nitric oxide synthase (eNOS), whereby the enzyme eNOS produces detrimental amount of superoxide anion  Atherosclerotic cardiovascular disease and vascular complications associated with risk factors such as diabetes mellitus, hypercholesterolemia, hypertension, aging, etc., remain the most important challenge for our society  and nitric oxide (NO⋅) are important signaling molecules involved in the regulation of vascular functions, including vascular relaxations, inflammatory responses, and cell proliferation . 2O2 which is either detoxified to H2O by peroxiredoxins, glutathione peroxidases, and catalase or metabolized to the powerful oxidant molecules such as hydroxyl radical (OH⋅), peroxynitrite , flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN), and tetrahydrobiopterin (BH4). Electrons from NADPH are transferred in trans from the carboxyl terminal reductase domain of one eNOS monomer, via the flavins FAD and FMN, to the heme in the amino-terminal oxygenase domain of the other monomer, where BH4, oxygen, and l-arginine are bound  that competes with l-arginine for eNOS binding  and arginase-II (Arg-II), which is encoded by two separate genes. The human Arg-I gene which maps to chromosome 6q23, encodes a 322 amino acid protein  as well as intracellular l-arginine concentration (0.05–0.2 mmol/L) far exceed the Km of eNOS (2–20 μmol/L)  even in the presence of high extracellular concentration of l-arginine (0.4 mmol/L)  does not have any benefits on vascular stiffness and left ventricular ejection fraction, but increases mortality .The underlying mechanisms of the detrimental effects of chronic ⋅ production is well documented  and peroxynitrite increase Arg-I gene expression in porcine coronary arterioles  is also implicated in regulation of arginase expression and activity in endothelial cells. p38MAPK is a member of the superfamily of MAPKs which serves as cellular a stress sensor for a variety of cellular stresses including hyperglycemia, oxidative stress, and inflammatory cytokines  and Arg-II in endothelial cells (Yepuri et al., /−− background show accelerated atherosclerosis (Vaisman et al., Arginase-II as therapeutic target in cardiovascular diseases has shown promising beneficial effects in genetic modified mouse models. Systemic deficiency of Arg-II reduces systemic and vascular inflammations in mice fed high cholesterol diet and high fat diet, and improves endothelial function in aging, reduces atherosclerosis, and improves insulin sensitivity and glucose homeostasis (Ming et al., It is however, interesting to notice that pharmacological agents that target arginase indirectly through blockade of signaling transduction pathways that regulate arginase gene expression or activity show beneficial effect on vascular functions. Early studies demonstrate that statins which inhibit arginase activity through inhibition of the small G protein or GTPase RhoA improves endothelial function (Ming et al., 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.


To compare the efficacy of fine-needle non-aspiration cytology (FNNAC) with that of fine-needle aspiration cytology (FNAC) of thyroid lesions.FNAC and FNNAC techniques were studied in 50 cases of thyroid lesions. All the needle-sampling procedures were done by a single operator. The samples were assessed cytologically and evaluated using five parameters, that is, background blood or clot, amount of cellular material, degree of cellular degeneration, and degree of cellular trauma and retention of appropriate architecture.Wilcoxon signed rank test was performed using SPSS14 software. Differences between all the individual parameters as observed in FNAC and FNNAC smears were insignificant.After evaluation of FNAC and FNNAC on the basis of these scores, greater numbers of diagnostically superior samples were obtained by FNNAC; however, by FNAC more number of diagnostically adequate smears were observed. The numbers of unsuitable smears were also more by FNNAC technique. In vascular organs, an alternative method of fine-needle non-aspiration cytology (FNNAC) also known as cytopuncture was developed in France in 1982 by Brifford et al.[The study population comprised all patients who presented with thyroid swellings at the Department of Pathology (cytology section) from May 2005 to April 2007. After a thorough clinical examination, all the patients were subjected to both FNAC and FNNAC. A total of 50 cases of thyroid lesions were included in this study. The procedure was explained to the patient and verbal consent was obtained prior to performing the procedure. The patients were subjected to FNAC and FNNAC using 23-gauge needles and 10-cc plastic syringes. All the procedures were performed by a single operator. FNAC or FNNAC sampling was carried out randomly with lesions, irrespective of consistency and size of lesions. Every slide was assessed without the prior knowledge of techniques utilized. The study was thus single blind and also prevented the observer bias. The smears were scored according to criteria using a predetermined scoring developed by Mair et al..A cumulative score between 0 and 10 points was obtained for each specimen which was then categorized into one of the following three categories:Category 1—(Score 0–2) Unsuitable for diagnosis.Category 2—(Score 3–6) Adequate for cytological diagnosis.Category 3—(Score 7–10) Diagnosis superior.P=0.05.The difference in the score for the individual parameter was assessed by Wilcoxon signed rank test using SPSS14 software. All the results were analyzed considering the statistical significance at a level of The non-aspiration technique yielded less diagnostically adequate but more diagnostically superior smears when compared with aspiration technique. A total of 19 cases were unsuitable for cytodiagnosis by non-aspiration as compared with 17 cases by aspiration technique .P value obtained by Wilcoxon signed rank test was not statistically significant in favour of non-aspiration sampling for any parameter. However, the average scores for each parameter favoured non-aspiration sampling than aspiration sampling [sampling .Non-aspiration sampling displayed more cellular material, less cellular trauma and degenerative changes, better retention of architecture and less likelihood of obscuring by blood. The average score per case was 6.04 by non-aspiration technique and was 5.90 by aspiration technique. FNAC, since its inception in 1847, has passed through two phases of initial scepticism and interim enthusiasm and has successfully reached the final stage of acceptance as identified by Orell in his aThe important advantage of FNNAC sampling is easy operation and absolute control over operating hand, especially for neck, breast, cutaneous or subcutaneous tissue. The FNNAet al.,[et al.[et al,[et al.[Results when compared for background blood contamination supported the non-aspiration technique Figures  and 2 buet al., the amouet al., and 4, b.,[et al. observed.,[et al. was greal.[et al, but the l.[et al, and 4 sil,[et al. but the l,[et al. and 4 wiet al.[For the five parameters studied objectively, there was no statistically significant difference observed between the two techniques, and a similar result has been found by Haddadi-Nezhad et al. Whereas et al.11et al.[et al.[In the present study, more diagnostically superior and less diagnostically adequate samples were obtained more by non-aspiration technique in comparison to aspiration technique. It was observed that the percentage of inadequate sampling was more with non-aspiration (38%) than with aspiration 34%) technique in contrast to the observations of Santos and Leiman[% techniqet al. With thel.[et al.Both the techniques have their own merits and demerits and neither is superior to the other. By combining both the techniques, better diagnostic accuracy can be achieved. However, FNNAC technique is easier to perform with better patient compliance.


Dear Editor in Chief,With a great interest, we read a recently published paper in Frontier in Endocrinology journal by Vuolo et al., entitled “Vitamin D and Cancer”  by b-galactosidase and sialidase of B and T cells, respectively, which can activate macrophage to phagocyte the neoplastic cells. Group-specific component protein (Gc) as DBP preserve vitamin D in body fluids and put it available for tissues. But an enzyme  produced by neoplastic cells deactivate and denaturize this factor and provide neoplastic cells to spread (Yamamoto and Naraparaju,


Arabidopsis and maize have identified a number of genes that are required for epidermal cell differentiation. However, the mechanism that specifies shoot epidermal cell fate during plant organogenesis remains largely unknown. Particularly, little is known regarding positional information that should restrict epidermal cell fate to the outermost cell layer of the developing organs. Recent studies suggested that certain members of the HD-ZIP class IV homeobox genes are possible master regulators of shoot epidermal cell fate. Here, we summarize the roles of the regulatory genes that are involved in epidermal cell fate specification and discuss the possible mechanisms that limit the expression and/or activity of the master transcriptional regulators to the outermost cell layer in plant shoots.Land plants have evolved a single layer of epidermal cells, which are characterized by mostly anticlinal cell division patterns, formation of a waterproof coat called cuticle, and unique cell types such as stomatal guard cells and trichomes. The shoot epidermis plays important roles not only to protect plants from dehydration and pathogens but also to ensure their proper organogenesis and growth control. Extensive molecular genetic studies in  The shoot epidermis is a single layer of surface cells that are morphologically characterized by anticlinal cell division patterns. The outer surface of the shoot epidermis is covered with a hydrophobic structure called a cuticle, which prevents water loss, pathogen attacks, and post-genital fusion of organs  was identified as an epidermis-specific homeobox gene that belongs to the HD-ZIP class IV family  after the embryos have undergone tangential cell divisions to generate outer protodermal cells and inner cells  caused severe phenotypes associated with defects in epidermal cell specification , a rice HD-ZIP class IV gene, was induced on the cut surface during callus regeneration  mutant, which develops multilayered epidermis by amplification of differentiated epidermal cells, expression of an HD-ZIP class IV gene was detected only in the outermost epidermal layer , which serve as precursor of cuticular wax  and zhoupi (zou) mutants are defective in cuticle formation in organs generated during embryogenesis . Plant embryos may require ALE1 and GSO1/GSO2-mediated signaling for efficient deposition of cuticle on the surface of the protodermal cells that develop in close physical contact with surrounding endosperm cells.crinkly4 (cr4) is a maize mutant with defects in the development of leaf epidermis and aleurone layer ] cause phenotypes defective in epidermal cell differentiation, lateral root initiation, and root initial cell maintenance  and TOADSTOOL2 (TOAD2) are leucine-rich repeat receptor-like kinases redundantly required for epidermal cell differentiation in the embryo  in the surface cells  is a calpain like cysteine protease that is conserved among land plants (Lid et al., DEK1 mRNA is expressed ubiquitously, suggesting that its activity is regulated post-translationally (Wang et al., Arabidopsis was not sufficient to upregulate the expression of ATML1 (Johnson et al., DEK1 activity affected cell division and growth also in internal tissues, suggesting that the action of DEK1 is not epidermis-specific (Johnson et al., dek1 may cause discontinuity and abortion of the epidermis (Johnson et al., Localization of some HD-ZIP class IV transcription factors was not limited to nuclei of heterologous cells (Zhang et al., in planta and it is possible that dimerization with other HD-ZIP class IV proteins changes the activity of ATML1 in a cell-type dependent manner, although ectopic expression of ATML1 alone was sufficient to induce epidermal cell fate in inner tissues (Takada et al., ATML1 was shown to heterodimerize with PDF2 atml1;pdf2 and DEK1 knockdown lines showed ectopic differentiation of mesophyll cells on the surfaces of leaves and cotyledons, respectively, (Abe et al., rpk1;toad2 embryos exhibited ectopic subepidermal marker expression in the outermost cell layer (Nodine et al., ATML1 decreased differentiation of green mesophyll cells in leaves (Takada et al., cr4 and dek1, which are defective in “surface” aleurone layer differentiation in the maize endosperm, cause ectopic differentiation of “inner” starchy endosperm cells on the surface of the endosperm (Becraft et al., Several lines of evidence show that acquisition of epidermal cell fate is associated with a loss of mesophyll or internal cell fate. First, epidermis-deficient Mesophyll cells possibly represent a primitive state of leaf cells, considering that ancestral aquatic algae are composed mainly of mesophyll-like cells. Land plants may have repressed mesophyll cell differentiation to evolve an epidermis on the surface. Evolutionary studies, including comparative genomics, may be useful for identifying molecular components that promote epidermal cell formation (Zalewski et al., Despite the extensive molecular genetic studies in model plants, positional signals that specify shoot epidermal cell fate remain unknown Figure . Most ofThe cuticle-bearing outermost cells should have distinct mechanical properties compared with inner cells. Moreover, cells located at the surface are unique, in that they are in constant contact with the environment. These unique properties could influence the differentiation of epidermal cells. Attempts to directly isolate epidermis-promoting biomolecules and to identify physical/environmental constraints influencing epidermal cell fate may shed new light on the issue.Shinobu Takada wrote the main manuscript text and Hiroyuki Iida prepared Figure 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.


With as many as 300,000 United States troops in Iraq and Afghanistan having suffered head injuries Miller, , traumat It is estimated that as many as 300,000 U.S. troops in Iraq and Afghanistan have suffered head injuries Miller, . In the Largely due to population-wide increases in life-span, AD is rapidly becoming the public health crisis of our time. There are currently over three million Americans afflicted with the disease, a figure that is projected to increase to nearly nine million Americans and over 100 million world-wide by 2050  peptides], neurofibrillary tangles (NFTs), and widespread loss of cortical neurons Selkoe, . AlthougThe term “TBI” encompasses a wide variety of traumas. In fact, any form of brain injury is broadly classified as a TBI. Nevertheless, brain traumas can grossly be divided into two categories: (1) closed head injuries (where a rapid deceleration or blow to the head causes brain damage) or (2) penetrating head injuries (caused by a foreign object piercing the skull). Closed head injuries can come in the form of skull fractures, brain contusions caused by brain-skull impact, hematomas, and diffuse axonal injuries brought on by shearing forces. Notably, closed head injuries associated with concussions from contact sports and shockwave blasts from improvised explosive devices have garnered much recent attention. TBIs may range from mild to severe, with about 75% of injuries coming in the form of concussions or other mild TBIs  has been demonstrated in the acute response to brain injury (Roberts et al., A very recent study examined survivors of a single TBI 1–47 years after the trauma, and reported that NFTs and Aβ pathology were present in approximately one-third of these patients. Such findings demonstrate the long-term consequences of a single TBI event (Johnson et al., As mentioned above, TBI is a strong epigenetic risk factor for development of AD later in life. Strikingly, several defining AD pathological hallmarks have been observed following TBI in patient brains and in numerous TBI animal models. In addition to neuronal and synaptic loss (Kotapka et al., Aβ deposits and widespread axonal Aβ accumulation have been found in patients' brains shortly after TBI (Roberts et al., de novo following brain trauma (Cribbs et al., In an attempt to clarify mechanisms of plaque appearance after brain trauma, several non-transgenic rodent and rabbit models have been utilized. While these animal models have proved useful to characterize axonal Aβ accumulation after TBI, wild-type rodents and rabbits did not manifest cerebral β-amyloid plaques. This is likely owed to the fact that these TBI animal models have relatively low abundance of brain endogenous Aβ species that do not reach a critical threshold for aggregation (Lewen et al., An important related concept is the idea of Aβ pathology “spreading.” Interestingly, studies from Mathias Jücker's laboratory and others have shown that intracerebral infusion of brain extracts containing aggregated Aβ can initiate Aβ deposition in brains of APP transgenic mice (Kane et al., TBI has also been shown to induce tauopathy. In that regard, it is important to note that a similar process has been described for spreading of NFTs by axonal transport after injection of abnormally folded tau filaments into a mouse model of cerebral amyloidosis (Clavaguera et al., In patients' brains as well as in experimental animal models, TBI has been associated with microglial activation (Carbonell and Grady, Depending on their activation state, microglia can be deleterious or beneficial in the context of cerebral amyloid deposition (Town et al., The development of clinically-relevant animal models is critically important to enable future study at the intersection of TBI and AD research. Animal models of AD fail to exhibit some of the key pathological earmarks of the human syndrome, even after significant brain injury (Uryu et al., By contrast, we have recently published a novel rat transgenic model of AD, line TgF344-AD. This transgenic line expresses mutant human APP and presenilin-1, which are each independent genetic causes of early-onset familial AD. Notably, this rat displays the full spectrum of human AD hallmarks, including cerebral amyloidosis, tauopathy, gliosis, and most importantly, large-scale apoptotic loss of neurons in cortical and hippocampal regions. Moreover, these animals display significant age-dependent cognitive disturbance (Cohen et al., The correlation between brain injury and neurodegenerative disease is now well-established (Szczygielski et al., These mechanistic investigations and the development of pre-clinical therapeutics will rely critically on a clearer understanding of both human pathologies. A key limiting factor is the large gap in our knowledge of the link between post-mortem observations in humans after TBI with animal model systems. Part of the uncertainty can be attributed to limitations inherent to experimental models of TBI. Therefore, it is expected that more precise modeling of pathological hallmarks in animal models will allow us to fill the knowledge gap. Specifically, it will be critical to develop models that accurately mimic the forces impacting the human brain under a variety of circumstances (Morales et al., 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.


Computed tomography revealed a marked decrease in the size of the metastases following three therapeutic courses, and no lung metastases or new lesions were detected following nine therapeutic courses. The response was declared clinically complete. The patient refused additional treatment following nine therapeutic courses, and there was no recurrence 36 months after the final course of therapy. This case demonstrates the efficacy of IRIS plus bevacizumab as a first-line combination therapy against lung metastases of rectal cancer.This report presents the case of a 72-year-old male who had undergone abdominoperineal resection following a diagnosis of lower rectal cancer with multiple lung metastases. Pathologically, the resected specimen exhibited advanced rectal cancer with regional lymphoid metastases and was classified as stage IV disease. S-1 and irinotecan (IRIS) plus bevacizumab combination therapy was used to treat the lung metastases following the surgery. S-1 (100 mg/body) was administered orally on days 1–14 of a 28-day cycle, and irinotecan (125 mg/m As a result of marked improvement in systemic chemotherapy against unresectable and/or recurrent colorectal cancer, the median survival time of patients with metastatic colorectal cancer has improved to >20 months with administration of fluorouracil (5-FU), irinotecan and oxaliplatin . Accordi2) and bevacizumab (7.5 mg/kg) were administered by intravenous infusion on days one and 15. Following three courses of therapy, the metastatic right lung tumor decreased in size to ~10 mm in diameter, and the left lung tumor had decreased in size to ~3 mm in diameter  with a diagnosis of rectal cancer. Patient medical history was otherwise unremarkable. On physical examination, all findings were unremarkable with the exception of slight pallor in the palpebral conjunctiva. Hematological investigations revealed anemia . Other laboratory tests and serum levels of carcinoembryonic antigen and carbohydrate antigen 19-9 were all within normal limits. Colonoscopy revealed the entire circumference of an elevated tumor with central depression and erosion at the lower rectum. A biopsy specimen from the tumor was indicative of a moderately differentiated adenocarcinoma. Abdominal computed tomography (CT) revealed thickening of the rectal wall with regional lymph node swelling but no liver metastasis. Chest CT revealed two metastatic lung tumors measuring 25 mm (in the middle lobe of the right lung) and 10 mm in diameter (in the lower lobe of the left lung) . A diagndiameter . Followidiameter . Followidiameter . The reset al (et al (et al (Multidisciplinary therapies, including systemic chemotherapy against unresectable and/or recurrent colorectal cancer, have improved, and FOLFOX, CapeOX and FOLFIRI combination chemotherapies plus bevacizumab or anti-EGFR monoclonal antibody can improve the survival times of patients. In the present case, the patient received IRIS plus bevacizumab combination therapy against lung metastases of rectal cancer. Regarding IRIS chemotherapy, Muro et al  reportedl (et al  and Tourl (et al  identifil (et al , IRIS iset al (et al (et al (The effectiveness of IRIS plus bevacizumab combination therapy as a first-line therapy for metastatic colorectal cancer has been reported. In a phase II study, Komatsu et al  reportedl (et al  reportedl (et al  reportedTreatment of lung metastases from colorectal cancer is a debated issue. The overall survival of patients with completely resectable lung metastases is better than that of patients with unresectable lung metastases –17. The In conclusion, IRIS plus bevacizumab combination therapy is well tolerated and efficacious as first-line chemotherapy for metastatic colorectal cancer. As a CV port is not required and patients can be released from the infusion pump while at home, IRIS plus bevacizumab combination therapy could contribute to improved quality of life in patients with metastatic colorectal cancer.


Normal aging is generally characterized by a slow decline of cognitive abilities albeit with marked individual differences. Several animal models have been studied to explore the molecular and cellular mechanisms underlying this phenomenon. The excitatory neurotransmitter glutamate and its receptors have been closely linked to spatial learning and hippocampus-dependent memory processes. For decades, ionotropic glutamate receptors have been known to play a critical role in synaptic plasticity, a form of adaptation regulating memory formation. Over the past 10 years, several groups have shown the importance of group 1 metabotropic glutamate receptor (mGluR) in successful cognitive aging. These G-protein-coupled receptors are enriched in the hippocampal formation and interact physically with other proteins in the membrane including glutamate ionotropic receptors. Synaptic plasticity is crucial to maintain cognitive abilities and long-term depression (LTD) induced by group 1 mGluR activation, which has been linked to memory in the aging brain. The translation and synthesis of proteins by mGluR-LTD modulate ionotropic receptor trafficking and expression of immediate early genes related to cognition. Fragile X syndrome, a genetic form of autism characterized by memory deficits, has been associated to mGluR receptor malfunction and aberrant activation of its downstream signaling pathways. Dysfunction of mGluR could also be involved in neurodegenerative disorders like Alzheimer’s disease (AD). Indeed, beta-amyloid, the main component of insoluble senile plaques and one of the hallmarks of AD, occludes mGluR-dependent LTD leading to diminished functional synapses. This review highlights recent findings regarding mGluR signaling, related synaptic plasticity, and their potential involvement in normal aging and neurological disorders. Over the next decades, a majority of developing countries will face one of the biggest challenges in history: accelerated aging of its populations. In human, aging is characterized by physical, psychological, and social changes. Interestingly, major individual differences are known to exist and low probability of disease and disability, high cognitive and physical capacity, and active engagement in life in general are associated to successful aging. Several animal models have been developed to investigate the processes behind this phenomenon  test was introduced in 1984, to study hippocampus-dependent memory in rats Morris, . CognitiEarly on, alterations in cholinergic neuronal activity were linked to memory impairments in the Long-Evans rat model  receptor subtypes , another form of synaptic plasticity has been closely related to memory formation -terminal domain of group 1 mGluR  isoform  and Alzheimer’s disease (AD). Results published on animal models of those diseases and clinical trials conducted with patients will thus help to understand the processes involved in the aging of the brain.FMR1 gene transcription which encodes the Fragile X mental retardation protein (FMRP; Pieretti et al., FMR1 KO mice and FXS patients could dysregulate synaptic plasticity and lead to memory deficits (Luscher and Huber, Group 1 mGluRs have been implicated in the development of the FXS (Dolen et al., FMR1 KO mice, the lack of this protein synthesis suppressor may unbalance cellular mechanisms and exaggerate mGluR-dependent signaling (Dolen and Bear, FMR1 KO mice brain, mGluR5 is associated more with Homer 1a stimulating agonist-independent signaling pathways than long Homer isoforms (Ronesi et al., FMR1 KO mice, mGluR-LTD formation does not require the acute stimulation of protein synthesis which is probably related to the constitutive overexpression of proteins implicated in LTD formation and maintenance to compensate for the lack of FMR1 (Hou et al., FMR1 KO mice, LTP priming is reduced by mGluR decoupling from protein synthesis (Auerbach and Bear, The stimulation of group 1 mGluR with the specific agonist DHPG normally induces FMR protein translation in dendrites (Weiler et al., Drosophila fly model (McBride et al., in vivo treatment with group 2 mGluR antagonists in these mice (Choi et al., FMR1 KO mice by reducing mGluR-LTD with a 50% reduction in mGluR5 expression (Dolen et al., The first demonstration of mGluR modulation rescuing phenotypes in FXS animal models was attained by pharmacologically rescuing social interaction and memory impairments in the FMR1 KO mice brain (D’Agata et al., Drosophila FMRP gene and presenilin, a protein mutated in the familial form of AD (McBride et al., 2+ from intracellular stores, it can also facilitate the hyperphosphorylation of tau by up-regulating protein kinase activities (Tsai et al., 2+ signaling, synaptotoxicity, and eventually synapse deterioration (Renner et al., A possible link between FXS and AD has been proposed since FMR protein binds to the amyloid precursor protein (APP; Sokol et al., After several decades of intense studies on processes involving ionotropic glutamate receptors in the aging brain, their metabotropic counterparts, and particularly post-synaptic group 1 mGluRs are becoming key targets to unravel synaptic plasticity processes underlying learning and memory. mGlu receptors, Homer proteins, and downstream signaling pathways likely all play critical roles in the maintenance of high cognitive abilities in old age. Further studies are thus highly warranted especially considering their apparent role in FXS and possibly AD.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The Athenian statesman and poet Solon (638–558 BC) believed that “no man can be happy,” being “wretched all those who contemplate the sun.” One century later, the lyric poet Pindar (c. 522–443 BC) wrote that there has never been a man “devoid of trouble” on earth, and there will never be one. In a recent Frontiers article, inspired by up-to date neuroscience evidence on the perception of pain, Lelard and colleagues investigated the interrelations between behavior and emotion in painful situations (Lelard et al., Unfortunately, we all know the unpleasant sensory and emotional experience of pain. What we can now explore scientifically in regards to pain has been complexly expressed through language and philosophy. The ancient Greeks used the word To determine how imagining oneself in painful situations might influence muscular and physiological responses, posturographic and physiological data were collected from participants viewing pictures of both painful and non-painful situations, while instructed to imagine themselves experiencing each situation. A displacement of center-of-pressure measure collected from a posturographic platform indicated that movement in the anteroposterior direction was shorter when subjects were presented with painful images. Responses of the tibialis anterior and soleus muscles, which have been associated with freezing responses in previous reports (Carpenter et al., Previous investigations have differentiated the “representational characters” that emerge while imagining a situation from the “instinctual characters” that arise in more automatic bodily responses, and have suggested that these may emerge from cortical vs. subcortical activity, respectively (Giummarra et al., The article by Lelard and colleagues suggests that emotional content is capable of changing muscular responses in specific configurations with ecological significance (Lelard et al., Additional questions may inform future research. For example, in what situations might emotionally-laden stimuli evoke responses different from those observed in this study—a muscular response indicative of fleeing rather than freezing? Would active suppression of empathy or perspective-taking for painful stimuli alter muscular responses? In addition, would suppression effectiveness differ for tasks with more visceral-response vs. representational-response evoking images? Possible experimental avenues for theoretical exploration that might support the implications of this study may exist in research examining individuals with unusual perspective-taking in healthy and pathological conditions—i.e., those with mirror-touch synesthesia (Banissy and Ward,


They have important microbicidal activities, however in inflammatory conditions they may secondarily attack surrounding tissues. Overproduction of reactive oxygen species, prolonged or excessive liberation of MPO and other effective yet also toxic substances from neutrophils may participate in disturbed apoptosis, intensify the inflammatory processes and result in serious human diseases. The inhibitory effect of quercetin on PMA stimulated SO generation in isolated human neutrophils was found to be dose-dependent, without affecting the activity of intact isolated neutrophils. At comparable conditions, quercetin was more potent in inhibiting MPO release than SO generation. Our results indicate that quercetin could support resolution of inflammation through decreased activity of neutrophils,  The concentration of 10 µmol/l significantly inhibited only MPO release by 36%. Our results are in agreement with findings of Nosal et al.,  and Pere (et al.  where quet al., et al., et al., 2+-ATPase (Horakova, et al., et al.,Quercetin is believed to protect human organism against several degenerative diseases (Hollman & Katan, Our study provided evidence supporting the potential beneficial effect of quercetin in diminishing tissue damage at the site of inflammation by inhibiting MPO release and by decreasing the generation of superoxide and the subsequently derived ROS.


The phenotype of DBA/2-mdx mice also seems to depend on the function of satellite cells. In this review, we summarize the methodology of direct isolation, characterization, and molecular regulation of satellite cells based on our results. The relationship between the regenerative capacity of satellite cells and progression of muscular disorders is also summarized. In the last part, we discuss application of the accumulating scientific information on satellite cells to treatment of patients with muscular disorders.Skeletal muscle has great regenerative capacity which is dependent on muscle stem cells, also known as satellite cells. A loss of satellite cells and/or their function impairs skeletal muscle regeneration and leads to a loss of skeletal muscle power; therefore, the molecular mechanisms for maintaining satellite cells in a quiescent and undifferentiated state are of great interest in skeletal muscle biology. Many studies have demonstrated proteins expressed by satellite cells, including Pax7, M-cadherin, Cxcr4, syndecan3/4, and c-met. To further characterize satellite cells, we established a method to directly isolate satellite cells using a monoclonal antibody, SM/C-2.6. Using SM/C-2.6 and microarrays, we measured the genes expressed in quiescent satellite cells and demonstrated that Hesr3 may complement Hesr1 in generating quiescent satellite cells. Although Hesr1- or Hesr3-single knockout mice show a normal skeletal muscle phenotype, including satellite cells, Hesr1/Hesr3-double knockout mice show a gradual decrease in the number of satellite cells and increase in regenerative defects dependent on satellite cell numbers. We also observed that a mouse's genetic background affects the regenerative capacity of its skeletal muscle and have established a line of DBA/2-background  One of the best-known examples of regeneration is the ability of newts to regenerate limbs and tails. This process was believed to utilize specialized multipotent cells that have the potential to produce all types of cells. The existence of this type of multipotent stem cells seemed to be the reason newts, but not humans, can regenerate limbs. However, a recent study of the axotl has suggested a different model  cells, and muscle-resident interstitial cells, that seemed to function as stem cells for muscle regeneration integrin β1(+)Cxcr4(+)CD34(+)CD45(−)Sca-1(−)Mac-1(−) fraction contained only myogenic cells  constitute a subgroup of the transforming growth factor (TGF)-β superfamily, and are known as myogenic differentiation regulators. Gamell et al. reported that Bmp2 induces phosphorylation of Akt and migration of C2C12  were upregulated (>5-fold) in the activated state in our microarray results. The most highly upregulated gene (334-fold) was As mentioned above, satellite cells occupy a unique location and do not express the myogenic determination gene MyoD. Therefore, we hypothesized that the genes responsible for maintaining satellite cells are specifically expressed in quiescent satellite cells in skeletal muscle. To isolate such genes, we also prepared non-myogenic cells from skeletal muscle for comparison with quiescent and activated satellite cells, and 63 genes were finally identified as “quiescence genes,” which are highly expressed in quiescent satellite cells but not in cultured myoblasts and non-myogenic cells in skeletal muscle , but these genes are also expressed in non-myogenic cells in skeletal muscle. Another discrepancy is the expression of Notch-related genes in quiescent satellite cells. We identified Notch signaling-related genes (Notch3 and HeyL/Hesr3) as the most highly expressed genes in quiescent satellite cells, although Pallafacchina's results did not show the importance of Notch signaling in quiescent satellite cells. However, recent studies have clearly demonstrated the essential roles of Notch signaling for maintaining quiescent satellite cells in vivo as well as developmental stages (see below). On the other hand, quiescent satellite cells share common features in our and Pallafacchina's results, for example, some cell adhesion molecules and transcriptional factors. Although our reports had not mentioned it, Pallafacchina et al. intriguingly found up-regulation of anti-oxidative genes in quiescent satellite cells. In fact, our original data also included ant-oxidative genes in quiescent-stage specific manner. These results imply the importance of oxidative stress in quiescent satellite cells. Farina et al. analyzed gene expressions of quiescent satellite cells, activated satellite cells , and proliferating myoblasts   and Hesr  are known as primary targets of Notch signaling , which leads to a decrease in BrdU uptake  leads to the generation of mononuclear cells that proliferate  is a well-known inherited muscular disorder, and the causative gene, mdx mouse (the correct nomenclature is C57BL/10-DMDmdx) is the most widely used model animal of DMD  show similar phenotypes to humans; loss of muscle weight, increased fibrosis, accumulation of adipocytes, and decreased muscle force  and demonstrated that mdx/mTR mice exhibit severe muscular dystrophy and a decrease in the loss of satellite cell proliferation  . Recently, Cappellari et al. reported that Dll4 and PDGFβ signals convert myogenic cells to pericyte-like cells without erasing their myogenic memory (Cappellari et al., Satellite cells undoubtedly have the best potential to produce new myofibers in vitro reduces their regenerative activity, as described above (Montarras et al., in vitro (Gilbert et al., in vivo regenerative potential (Parker et al., Another problem is the difficulty of obtaining large numbers of satellite cells from a donor. In addition, expansion of satellite cells Another source of cells for therapy for muscle disorders is iPS cell-derived myogenic cells (Mizuno et al., mdx phenotype (Le Grand et al., The physiological self-renewal mechanism of satellite cells during regeneration must also be revealed. Some evidences have indicated an asymmetrical model of self-renewal of satellite cells (Conboy and Rando, In addition, another mechanism must be understood for the successful cell transplantation for muscle disorders. To date, we have not paid attention to the fusion process between transplanted cells and myofibers. When donor cells are transplanted, they have to cross the basal lamina and fuse with myofibers. Horsley et al. indicated the myotube-myoblast fusion process requires IL4/IL-4R signaling mediated by NFATc2 (Horsley et al., For 10 years, the isolation, characterization, and molecular regulation of satellite cells have been studied, and accumulating evidence is starting to reveal the molecular mechanisms for maintaining the satellite cell pool. In addition, recent studies are beginning to identify the roles of satellite cells in several different disorders. For instance, McCarthy et al. indicated that acute hypertrophy is not dependent on satellite cells (McCarthy et al., The generation of iPS cells has assisted the cell therapy approach to many disorders including muscular dystrophy (Takahashi and Yamanaka, Skeletal muscle is indispensable for motility. In addition, skeletal muscle has the potential to control other tissues. For instance, Bostrom et al. showed that a skeletal muscle-derived cytokine, irisin, converts white fat cells to brown fat-like cells (Bostrom et al., 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.


A decline in skeletal muscle mass and function with aging is well recognized, but remains poorly characterized at the molecular level. Here, we report for the first time a genome-wide study of DNA methylation dynamics in skeletal muscle of healthy male individuals during normal human aging. We predominantly observed hypermethylation throughout the genome within the aged group as compared to the young subjects. Differentially methylated CpG (dmCpG) nucleotides tend to arise intragenically and are underrepresented in promoters and are overrepresented in the middle and 3′ end of genes. The intragenic methylation changes are overrepresented in genes that guide the formation of the junction of the motor neuron and myofibers. We report a low level of correlation of gene expression from previous studies of aged muscle with our current analysis of DNA methylation status. For those genes that had both changes in methylation and gene expression with age, we observed a reverse correlation, with the exception of intragenic hypermethylated genes that were correlated with an increased gene expression. We suggest that a minimal number of dmCpG sites or select sites are required to be altered in order to correlate with gene expression changes. Finally, we identified 500 dmCpG sites that perform well in discriminating young from old samples. Our findings highlight epigenetic links between aging postmitotic skeletal muscle and DNA methylation. We found an overall global trend of genomic hypermethylation in human skeletal muscle between the young and old groups Fig. . In ordeet al., et al., et al., CpG islands (CGI) are present in 60% of gene promoters in the 450 K chip, and methylation deregulation in CGI overlapping the promoter has often been linked with cancer  negatively correlates with the corresponding promoter methylation level  was the transcription factor CTCF . This suggests that the dmCpG sites we identified within the aged group are relevant to muscle tissue. There were 54 enriched terms with a P < 0.01 (Table P = 2.05E-07) and axon guidance (P = 2.11E-07), which may be related to muscle innervation and aging. A decline of function in the neuromuscular junction has long been thought to contribute to the decline of muscle mass with age , cytoskeleton (P = 0.0003), and cytoskeleton organization (P = 0.001) are all linked to cytoskeleton function, and the latter plays an important role in proper muscle contractile function , homophilic cell adhesion (P = 8.43E-08), and cell adhesion (P = 1.70E-05), with cell adhesion being associated with fiber degeneration  (Table P = 6.16E-10) with 50 of 216 pathway members (23.1%) differentially methylated in at least one intragenic site .A recent report utilized genes identified as having dmCpGs with age to examine stem cell differentiation pathways  occurs with aging and is a major contributory factor in sarcopenia (Doherty et al., et al., et al., et al., et al., In aging human skeletal muscle, we identified a strong preference for the dmCpG sites to localize within a gene, and in the central and 3′ end regions of genes, but surprisingly no preference for the promoter. Some studies have suggested that intragenic DNA methylation could regulate alternative splicing (Sati et al., A potential issue with the biopsy procedure used in our study is tissue heterogeneity between biopsies, which may impact the resultant molecular signatures. There are other cell types in muscle that might contaminate muscle-specific methylation signatures. Such sample variation may influence the results. However, it was previously demonstrated that there is remarkably little technical variation between discrete biopsies sampled from the right and left leg of the same individual with regard to fiber type differences, gene expression as assessed by microarray, fiber size, or even strength (Tarnopolsky et al., et al., In summary, we describe for the first time a genome-wide DNA methylation survey of a postmitotic tissue between young and old healthy human skeletal muscle. We also determined that there were several hundred robust dmCpGs that discriminated younger males from older males. Ontology analysis of the terms associated with dmCpGs provided several interesting hints of a potential role for epigenetic changes in the neuromuscular junction during aging. Resistance exercise can reverse specific aspects of the gene expression changes in aging to more youthful levels (Melov All methods and experimental procedures are available in online supplement.