Primary biliary cirrhosis (PBC) is a chronic cholestatic liver disease characterized by the immunological destruction of intralobular bile ducts and serum anti-mitochondrial antibodies (AMA). Based upon previous work of oral tolerance and autoimmunity, we hypothesized that feeding the mitochondrial autoantigens of PBC would alter the clinical course and the level of antimitochondrial antibodies. The bovine pyruvate dehydrogenase complex (PDC) was purified and 5 mg fed in gelatin capsules to 6 patients with early stage PBC for 6 months. Antimitochondrial antibodies and liver biochemistries were measured at every 3 months for 12 months. The clinical trial was completed for all patients except for 1 who showed deterioration of pre-existing skin rash during treatment, which disappeared within 2 weeks after treatment was discontinued. However, after 1 year, neither the titers of AMAs nor liver biochemistries were significantly changed by this treatment. This is the first trial to test the efficacy of oral tolerance induction in PBC. However, the data, which limited in scope, did not demonstrate efficacy and further highlights the difficulties in showing continuing evidence of tolerance induction in autoimmunity.


The immunofluorescent reaction of peripheral blood cells from 10 multiple myeloma, 10 Hodgkin's and 11 lymphosarcoma cases with antiviral (Rauscher) murine leukaemia (AMR) and antihuman stem cell leukaemia plasma (AHS) antisera was studied. Cells from 25 of these patients were reactive at least once with AMR and AHS or AHS alone. Absorption studies suggested that this cytoplasmic immunofluorescent reaction involved cellular isoantigens. Serial studies on multiple myeloma, Hodgkin's and lymphosarcoma cases with significant fluorescence, showed that the fluorescent cell count variation was correlated with the presence of active disease.


Insulin resistance has been extensively investigated during the past decade because of its proposed role in initiating a cluster of cardiovascular risk factors including hypertension. Insulin resistance is an inherited genetic trait that precedes hypertension in Dahl salt-sensitive (S) rats, and is not present in Dahl salt-resistant (R) rats. Owing to the co-existence of insulin resistance and salt sensitivity of blood pressure in Dahl S, but not R rats, Dahl S rats are used to elucidate the role of dietary salt as a potential link in exacerbating both phenotypes . In light of available data, examining the impact of dietary salt on insulin resistance in Dahl S rats in terms of salt concentration and duration of exposure helps answer the following question: What percentage of dietary salt and for what duration of exposure would we expect an enhanced insulin resistance in Dahl S rats? This commentary gathers all available research done on insulin resistance in Dahl S rats in an attempt to unravel dietary salt contribution to insulin resistance in Dahl S rats. Over the past decades, insulin resistance and compensatory hyperinsulinemia have been proposed as the common disorder underlying a constellation of risk factors  referred to as the metabolic syndrome or syndrome X . InsulinTo be able to address the above questions, Dahl S rats have been examined at various stages (weanlings and fully developed hypertensive rats) and with various dietary salt regimens . Weanling Dahl S rats when examined before developing hypertension and prior to sodium loading were found to be significantly insulin resistant as evidenced by their higher plasma insulin levels compared to weanling Dahl R rats . This imKnowing that Dahl S rats are genetically insulin resistant as evidenced by the presence of insulin resistance in weanling Dahl S rats as well as in fully hypertensive Dahl S rats, it's time to address our third and fourth questions which inquire about the role of salt in insulin resistance and whether a specific salt concentration and/or duration of exposure is required for salt to play a role in insulin resistance in Dahl S rats. Controversial reports in the literature have been documented with some studies proposing no effect of dietary salt in advancing insulin resistance and others proposing a significant role of salt in suppressing insulin signaling and precipitating insulin resistance in Dahl S rats. That's why, it is important to analyze all available data in terms of salt concentration and duration of exposure. Salt loading in Dahl S rats did not further increase their insulin resistance when fed 3% NaCl for 1 week , or 8% NNot only did 4 weeks of high salt diet of 8% NaCl exaggerated insulin resistance ,6and conSalt-induced oxidative stress and inflammation might be the mechanism by which high dietary salt concentration of 8% suppresses insulin signaling and precipitates insulin resistance in Dahl S rats. Inflammatory mediators enhanced by high dietary salt include the tumor necrosis factor-alpha  or JNK might possibly interfere with normal insulin signaling via a number of mechanisms including: i) disruption of insulin-induced insulin receptor substrate-1 (IRS-1) and phosphoinositide-3 kinase (PI3K) phosphorylation and cellular distribution, ii) reduction in glucose transporter (GLUT4) expression levels, or iii) reduction in GLUT4 translocation to the plasma membrane -11. SuppWhile one would expect suppressed insulin signaling in Dahl S rats, Ogihara et al., reported that Dahl S rats had an enhanced insulin signaling manifested by an enhanced tyrosine phosphorylation of the insulin receptor, insulin receptor substrates, activation of PI3K and phosphorylation of Akt compared to Dahl R rats. These effects could be simply attributed to the fact that the authors starved the rats for 12 hours prior to the experiment. It is well documented that 12-hour of fasting upregulated hepatic insulin receptors in Sprague Dawley rats (from which Dahl rats were derived)  Insulin resistance in Dahl S rats precedes hypertension, 2) Insulin resistance in Dahl S rats is significantly enhanced after 4 weeks of 8% NaCl, which represents the threshold for a salt-induced suppression in the insulin signaling pathway, 3) Salt-induced insulin resistance in Dahl S rats (after 4 weeks of 8% NaCl) is accompanied by an enhanced oxidative stress and inflammatory responses that is manifested by increased levels of TNF-alpha and JNK proteins, 4) Increased TNF-alpha and JNK levels might disrupt the insulin signaling pathway at the level of the insulin receptor substrates' association with either upstream or downstream molecules in the insulin pathway or via suppressing GLUT4 expression and/or translocation. Finally, it is worth mentioning that the promoter regions of the insulin receptor substrates –1, and –2 comprises putative sodium-responsive elements that might be acted upon by excess salt to suppress their corresponding activity, expression and/or function ,17. Muta


The assessed lack of validated reference genes emphasizes the importance of a systematic study for their identification. For selecting candidate reference genes we have developed a simple The expression stability of 32 genes was assessed by qRT-PCR using a set of cDNAs from 24 different plant samples, which included different tissues, developmental stages and temperature stresses. The selected sequences included 12 well-known HKGs representing different functional classes and 20 genes novel with reference to the normalization issue. The expression stability of the 32 candidate genes was tested by the computer programs geNorm and NormFinder using five different data-sets. Some discrepancies were detected in the ranking of the candidate reference genes, but there was substantial agreement between the groups of genes with the most and least stable expression. Three new identified reference genes appear more effective than the well-known and frequently used HKGs to normalize gene expression in wheat. Finally, the expression study of a gene encoding a PDI-like protein showed that its correct evaluation relies on the adoption of suitable normalization genes and can be negatively affected by the use of traditional HKGs with unstable expression, such as actin and α-tubulin.The present research represents the first wide screening aimed to the identification of reference genes and of the corresponding primer pairs specifically designed for gene expression studies in wheat, in particular for qRT-PCR analyses. Several of the new identified reference genes outperformed the traditional HKGs in terms of expression stability under all the tested conditions. The new reference genes will enable more accurate normalization and quantification of gene expression in wheat and will be helpful for designing primer pairs targeting orthologous genes in other plant species. Arabidopsis (using the large public collection of data from Affymetrix GeneChip experiments) -1) × 100 -1) × 100, where m is the slope of linear regression model fitted over log-transformed data of the input cDNA concentration versus Ct values according to the linear equation y = m*log(x) + b. Moreover, the Mx3000PTM software system computed the R2 value (Coefficient of determination), which represents an indicator of the quality of the fit of the standard curve to the plotted data points. The R2 values calculated for all the standard curves in each of the two biological replicates were higher than 0.994, indicating a strong linear relationship between the detected Ct values and the correspondent relative amount of template in all the amplification reactions  from pooled cDNA. By plotting the Ct values of five different data-sets: 1) all 24 samples including tissues, developmental stages and temperature treatments; 2) 18 samples relative to different tissues and developmental stages; 3) six samples consisting of two temperature treatments (4°C and 33°C), each for 24 and 48 h and their controls; 4) six samples relative to the single floral organs from fully emerged spikes; 5) six samples referring to vegetative tissues and developmental stages . These data sets were used to evaluate the expression stability of the genes belonging to the four families by 18 primer pairs, 14 of them targeting specific gene transcripts of the gene families (five each for actin and α-tubulin and two each for ADP-ribosylation factor and translation elongation factor) and four primer pairs which amplified multiple transcripts for each of the four gene families . Table t differences (ΔCt = Ctmax-Ctmin) obtained by the 18 primer pairs for each of the five data-sets considered, whereas the complete statistics obtained by ANOVA analysis are reported in Additional file t values gives a straightforward indication of the expression stability of a particular gene; most genes with the lowest CV values showed also the lowest values of Ct range (ΔCt) and MST (Mean Square of Treatments)  and 13 lowly expressed genes, with mean Ct values above this value , Ta2776 (RNase L inhibitor-like protein), Ta54448 and Ta54733 (Proteins of unknown function), and Ta44405 (β-tubulin), with mean Ct values between 26 and 24 cycles  and moderately expressed (10 genes with more than 300 EST). Of the 13 reference genes with low Ct values, only three including more than 300 ESTs  were considered moderately expressed in UniGene, whereas the expression level of the remaining 10 genes was low (<300/>100 ESTs) or very low (<100 ESTs). In our opinion, these data indicate a substantial agreement between the gene expression levels evaluated by qRT-PCR analyses and those estimated by the number of ESTs included in each corresponding UniGene cluster. Ta54963 (GABA-receptor associated protein), Ta54238 (Ubiquitin-conjugating enzyme), Ta35497 (Cyclophilin), Ta53967 (Vacuolar ATP synthase 16 kDa proteolipid subunit) exhibited the lowest gene expression variation (less than 3 cycles), while Ta44405 (β-tubulin), Ta38797 (Histone) and Ta1698 (NADP-isocitrate dehydrogenase) showed the most variable expression (more than 5 cycles).In order to avoid possible errors due to the interaction of co-regulated genes, further analyses were carried out on 32 genes involved in distinct biological processes and metabolic pathways; they comprised 28 genes selected on the basis of their uniform expression detected in UniGene and TIGR and a single gene for each of the four gene families. The expression levels could be grouped into two arbitrary categories: 19 highly expressed genes, whose mean CAs discussed in the previous paragraph, it was evident that none of the 32 selected candidate reference genes showed a constant expression under all the tested conditions  in wheat. Hence, as expected, it was not possible to find an all purpose reference gene, and it would be necessary to select the most suitable one for normalizing gene expression considering the peculiar and specific experimental conditions. The relationships between expression stability of the 32 candidate genes and different types of tissues and/or experimental conditions were tested by running the programs geNorm  and Normrd, Cell division control protein, AAA-superfamily of ATPases), Ta2291 , Ta53899 , Ta4045 , Ta2776 , Ta53967 , Ta54963  and Ta53919  , Ta2291 , Ta54238 , Ta54948 , Ta2776 , Ta30797 , Ta54227  and Ta4045  , Ta54227 , Ta54238 , Ta53891 , Ta54447 , Ta2291 , Ta53919  and Ta55512 , whereas Ta1698 (NADP-isocitrate dehydrogenase) and Ta44405 (β-tubulin), with M values of 0.63 and 0.59, were the least stable genes  tested in our study are plotted in figure se) Fig. . The ranse) Fig. . The ranse) Fig. , and theThe high expression stability of a gene indicates that the use of a single internal control is appropriate, but some studies require the normalization based on two or more stable internal control genes. The geNorm program, in addition to the gene stability measure M, computes a normalization factor (NF) and assesses the optimal number of reference genes for generating that factor. The pairwise variations Vn/Vn+1 between two sequential normalization factors (NFn and NFn+1) are used to determine the necessity of adding the next more stable control gene for reliable normalization. Pairwise variations were calculated using geNorm for each data set to determine the optimal number of internal control genes for normalization Fig. . In all nd, ADP-ribosylation factor), Ta2776 , Ta35284 , Ta53919 , Ta53967 , Ta54171 , Ta54963 , Ta54733  and Ta4045  ; the two least stable genes were Ta44405 (β-tubulin) and Ta38797 (Histone), with stability values of 0.150 and 0.198 respectively, the same identified as the worst genes by geNorm. The best combination of two genes suggested by NormFinder was that of Ta54227 and Ta2291, which improved the stability value to 0.030, indicating a more reliable normalization than that based on single genes. Similar results were obtained for 18 samples including tissues/organs at different developmental stages of wheat, with the lowest stability value (0.032) for Ta54227 (Table NormFinder (the second adopted program based on a model approach) ranks the best candidate references genes according to their minimal combined inter- and intra-group variation of expression. The results obtained by NormFinder analysis for the five data sets are shown in Table The stability ranking of the reference genes across the five analysed data sets obtained by geNorm and NormFinder was not identical for the genes with the most stable expression Table . In partThe expression levels of the target gene TaPDIL1-1 were used as an example to show the effect of using different normalization genes on estimating its relative expression. TaPDIL1-1 encodes the typical PDI (Protein Disulfide Isomerase) which accomplishes several metabolic functions, the most important consisting in disulfide bond formation and isomerization during the folding of secretory proteins ,40. PrevNormalization of the expression of TaPDIL-1-1 using the reference genes selected by the three methods based on geNorm and NormFinder programmes did not detect significant differences in the relative expression of the gene of interest in all the four analysed data sets Figs.  and 6. AArabidopsis to identify new reference genes, which has been based on the data furnished by the microarray datasets [in silico method exploiting UniGene and TIGR, two publicly available databases of wheat sequences enclosing non-redundant EST-indices. The method consists of two steps, each exploiting the peculiar features of these wheat-specific databases. Basically, an effective reference gene has to fulfil three main requirements: 1) moderate to high level of expression ; 2) expression in several tissues and developmental/physiological states; 3) low variation of gene expression across different tissues and developmental, physiological and environmental conditions [T. aestivum UniGene database (see Methods), which were accomplished by 177 out of the 42,256 sequence clusters therein contained. The second step consisted in the scanning of the TIGR wheat gene index database (TaGI version 11) for the identification of the TC (Tentative Consensus) or group of TCs corresponding to each of the 177 clusters previously selected in UniGene. The identification of TCs was important because these outputs are more effective for gene functional annotation and for designing gene specific qRT-PCR primers. The frequency  of each TC or group of TCs linked to the 177 selected UniGene clusters was determined and was used to pinpoint the most suitable candidate reference genes to be included in a preliminary screening.To the best of our knowledge, the present research represents the first effort aimed to the identification and systematic comparison of a large number of potential reference genes and of the corresponding primer pairs specifically designed for gene expression studies in wheat, in particular for qRT-PCR analyses. As mentioned in the "Introduction", so far most studies on gene expression in wheat have been carried out using single reference genes, without any preliminary validation of their expression stability  has been one of the most suitable reference genes in a set of tissue samples in rice, whereas the expression of UBQ10 was unstable [ACT2/7 was stably expressed, whereas ACT11 showed variable profiling [Among the 177 ol genes ,20,52. Tunstable . A similt values [Since the search for suitable reference genes is both time-consuming and cost intensive, several computer programs based on different statistical approaches have been developed to assist the selection of a reference gene among several HKGs. The software geNorm has been reported in 2002  and has t values . Accordit values , this apt values ,56, or tt values . The chot values ,57,58. St values  and Normt values  and compAs discussed by different authors ,57-59, sThe mathematical model of gene expression used by NormFinder enables to estimate both the overall variation of the candidate normalization genes and the variation between subgroups of the sample set. This approach has been reported to perform in a more robust manner than geNorm and has been shown to be less sensitive to the presence of co-regulated genes , therefot 24.79), whereas that of Ta2291 (mean Ct 21.97) and of Ta54227 (mean Ct 21.56) is high, they are suitable as reference genes in a large data set of wheat samples, but also for the normalization of target genes with contrasting expression levels.Expression stability analyses showed that the most stable reference gene identified by NormFinder and geNorm was not the same for the five data sets analysed Table . Even thth among the 32 candidate reference genes , α-tubulin (Ta25534), β-tubulin (Ta44405), Ubiquitin (Ta50503), GAPDH (Ta30768), Cyclophilin (Ta35497), Translation elongation factor (Ta53964), Translation initiation factor (Ta54280), Ribosomal protein (Ta27771) and Histone (Ta38797), in fact most of them classified as the least stable genes both by geNorm and NormFinder in the five data sets analysed , Ta2291 (ADP-ribosylation factor) and Ta2776 (RNase L inhibitor-like protein) resulted the best and most stable reference genes to normalize gene expression in different tissue and development stages of wheat; they outperformed all traditional HKGs such as α-tubulin (Ta25534), β-tubulin (Ta44405), Ubiquitin (Ta50503), Actin (Ta54825), GAPDH (Ta30768), Ribosomal protein (Ta27771) and Histone (Ta38797). Although in our experimental conditions a single reference gene was adequate to normalize the qRT-PCR data, the use of normalization factors generated from the expression data of two or three most stable genes should be considered in future studies to improve the accuracy and reliability of expression analysis. The set of qRT-PCR specific primers for the novel and more stable reference genes developed in this study will enable more accurate normalization and quantification of gene expression in wheat. Moreover, the design of suitable primer pairs for their orthologous genes could contribute to the development of reference genes with stable expression in other plant species.In the present research we compared 20 novel candidate reference genes and 12 commonly used HKGs suitable for gene expression normalization in wheat. Both new and traditional reference gene sequences were fetched by an qRT-PCR: quantitative Real-time RT-PCR; HKG: Housekeeping gene; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; RT: Reverse transcriptase; EST: expressed sequence tag; TIGR: The Institute for Genome Research; PDI: Protein disulphide isomerase; TPM: transcripts per million; TC: tentative consensus sequence; Mv: mean value; V: variance; SD: standard deviation; CV: coefficient of variation; SSH: Suppression Subtractive Hybridization; CS: Chinese Spring; TEF-1α: Translation Elongation Factor-1α; ADP-RF: ADP-ribosylation factor; GO: Gene Ontology; ANOVA: analysis of variance; MST: mean square of treatments; MSE: mean square errors; NF: normalization factor; NCBI: National Center for Biotechnology Information.ARP performed all the sample preparations and experimental procedures and participated in data analysis. MC supervised the study, performed data analysis and draft the manuscript. OAT performed data analysis, revised the manuscript critically and participated in tables and figures drawing. EP revised critically the manuscript and gave financial support to the study. All authors read and approved the final manuscript.List of articles reporting the application of reference genes for qRT-PCR normalization in wheat. The list was obtained by a PubMed search from January 1996 to March 2008 and includes 26 articles reporting 16 reference genes.Click here for fileRT-PCR primers used in this research. Table showing UniGene clusters and linked TC sequences of the reference genes used in qRT-PCR analysis, their primer sequences and characteristics of the corresponding amplicons.Click here for fileUniGene clusters and corresponding TC sequences of 177 selected candidate reference genes of wheat. The file reports the transcription profiles of the genes corresponding to the UniGene clusters in six wheat tissues  calculated as a value representing the number of transcripts per million (TPM). Mean value (Mv), Variance (V), Standard deviation (SD) and Coefficient of Variation (CV) were determined on the expression values (TPM) of the 177 Unigene clusters in the six wheat tissues. The highest and lowest expression values, the relative expression (percentage) in the six wheat tissues and the number of ESTs included in each UniGene cluster are also reported. The frequency  was determined for each TC or group of TCs linked to the 177 selected UniGene clusters.Click here for fileClassification of the 177 selected candidate reference genes of wheat on the basis of their putative functions. File showing the UniGene clusters and corresponding TC sequences of the 177 selected candidate reference genes of wheat and their putative functional annotation determined on the basis of their orthologous genes of rice and relative GO annotations.Click here for fileLevel of expression of the 177 selected candidate reference genes determined on the basis of the number of EST sequences included in each UniGene cluster. File showing UniGene clusters and corresponding TC sequences of the 177 selected candidate reference genes of wheat sorted in ascending order on the basis of the number of EST sequences included in each UniGene cluster.Click here for fileUniGene clusters and corresponding TC sequences of ten genes of the actin and α-tubulin families. Gene transcription profile of each UniGene cluster in six wheat tissues  is reported as a value representing the number of transcripts per million (TPM). Mean value (Mv), Variance (V), Standard deviation (SD) and Coefficient of Variation (CV) were calculated on the expression values (TPM) of the ten Unigene clusters in the six wheat tissues. The highest and lowest expression values, the relative expression (percentage) in the six wheat tissues and the number of ESTs included in each UniGene cluster are also reported. The frequency  was determined for each TC or group of TCs linked to the 177 selected UniGene clusters.Click here for filewcor14 and TaHSP101B.Expression analysis by RT-PCR of the genes  Agarose gels of RT-PCR products of wcor14 and TaHSP101B genes after 35 PCR cycles in six samples consisting of two temperature treatments (4°C and 33°C) for 24 and 48 h and their controls . The transcripts of the constitutively expressed gene encoding actin (UniGene cluster Ta54825) were amplified as control. M = part of the DNA molecular weight marker XIV (Roche), the most intense band is 500 bp in length.Click here for fileSpecificity of qRT-PCR amplification. Figures A1–A3 report the dissociation curves of 42 selected reference genes showing single peaks, they were obtained from three technical replicates of 18 cDNA pools representing different tissues and developmental stages of wheat. Figures B1–B3: Agarose gels (2%) showing amplification of a specific PCR product of the expected size for each candidate reference gene.Click here for filet values for 46 primer pairs and 24 tissue samples used in this research.PCR efficiency and C The file shows PCR efficiency and Ct values of two biological replicates performed on 24 tissue samples; three technical replicates were carried out for each of the 46 primer pairs. The 24 tissue sample comprised: 1) Roots from plants with single shoot and three leaves unfolded ; 2) Shoots1 = single shoots and leaves from plants at Feekes scale 1.3; 3) Shoots2 = shoots at the beginning of tillering ; 4) Shoots3 = shoots from plants with formed tillers ; 5) Shoots4 = shoots at the beginning of erect growth ; 6) flag leaves at booting stage ; 7) stems at booting stage ; 8–10) Spikes1–3 = spikes collected at intervals of 10–12 days ; 11–16) single floral organs  from fully emerged spikes ; 17–18) Seeds 1–2 = seeds collected 15 (medium milk stage) and 30 (hard dough stage) days after anthesis; 18–24) six samples  consisting of two temperature treatments (4°C and 33°C) for 24 and 48 h and their controls .Click here for filet values obtained in five different data-set. Expression stability of the genes belonging to four families  evaluated by statistical analysis of raw CANOVA analysis and statistics determined on raw Ct values obtained by 18 primer pairs which amplified either specific or multiple (m) gene transcripts of four gene families  in five different data-sets: 1) all 24 samples including tissues, developmental stages and temperature treatments; 2) 18 samples relative to different tissues and developmental stages; 3) six samples consisting of two temperature treatments (4°C and 33°C), each for 24 and 48 h and their controls; 4) six samples relative to the single floral organs from fully emerged spikes; 5) six samples referring to vegetative tissues and developmental stages .Click here for fileRanking of the expression stability of the reference genes as calculated by geNorm in four data sets. geNorm output charts of M values for the 32 selected reference genes in four data sets: (A) = 18 tissues and developmental stages; (B) = six samples consisting of two temperature treatments (4°C and 33°C) for 24 and 48 h and their controls; (C) = six floral organs from fully emerged spikes; (D) = six vegetative tissues and developmental stages .Click here for fileDetermination of the optimal number of control genes for accurate normalization calculated on the basis of pair-wise variation (V) analysis in four data sets. geNorm output charts of V values for the 32 selected reference genes in four data sets: (A) = 18 tissues and developmental stages; (B) = six samples consisting of two temperature treatments (4°C and 33°C) for 24 and 48 h and their controls; (C) = six floral organs from fully emerged spikes; (D) = six vegetative tissues and developmental stages .Click here for file


Two hundred and eleven patients with node-positive stage II and III breast cancer were treated with eight cycles of adjuvant chemotherapy comprising cyclophosphamide, doxorubicin and oral ftorafur (CAFt), with and without tamoxifen. All patients had undergone radical surgery, and 148 patients were treated with post-operative radiotherapy in two randomized studies. The impact of haematological toxicity of CAFt on distant disease-free (DDFS) and overall survival (OS) was recorded. Dose intensity of all given cycles (DI), dose intensity of the two initial cycles (DI2) and total dose (TD) were calculated separately for all chemotherapy drugs and were correlated with DDFS and OS. Patients with a lower leucocyte nadir during the chemotherapy had significantly better DDFS and OS (P = 0.01 and 0.04 respectively). Dose intensity of the two first cycles also correlated significantly with DDFS (P = 0.05) in univariate but not in multivariate analysis, while the leucocyte nadir retained its prognostic value. These results indicate that the leucocyte nadir during the adjuvant chemotherapy is a biological marker of chemotherapy efficacy; this presents the possibility of establishing an optimal dose intensity for each patient. The initial dose intensity of adjuvant chemotherapy also seems to be important in assuring the optimal effect of adjuvant chemotherapy.


Loss of mismatch repair proteins  or overexpression of nucleotide excision repair proteins (ERCC1) was not detrimental for the cellular sensitivity to BBR 3464. Moreover, interesting differences in the kinetics of formation and removal of DNA lesions at the single-gene (N-


Trans-[PtCl2(isopropylamine)] is a novel trans-platinum compound that shows cytotoxic activity in several cisplatin resistant cell lines. The aim of this paper was to analyse, by means ofmolecular cell biology techniques and total reflection X-ray fluorescence (TXRF), the cytotoxic activity, theinduction of apoptosis, the cellular uptake and the DNA binding of trans-[PtCl2(isopropylamine)] in the cisplatin resistant cell line A2780cisR. The results show that this drug is more cytotoxic and induces a higher amount of apoptotic cells than cisplatin in A2780cisR cells. However,the intracellular accumulation and extent of binding to DNA of trans-[PtCl2(isopropylamine)] is lower than that of cis-DDP. Moreover, trans-[PtCl2(isopropylaminae)] is partially inactivated by intracellular levels of glulathione. The resultsuggest that circumvention of ciplatin resistance by trans-[PtCl2(isopropylamine)] in A2780cisR cells might be related with the ability of this drug to induce apoptosis.


To investigate the underlying mechanisms of carcinogenesis, we have developed a technique to determine the frequency of genetic changes in prostatic carcinoma tissue. We have demonstrated that at a ratio of between 1:4 and 1:9 mutant-normal alleles, the signal from a mutant TP53 allele is not apparent after polymerase chain reaction (PCR) amplification and further direct sequencing or single-strand conformation polymorphism (SSCP) analysis. To bypass this problem, which is inherent in the heterogeneity of the prostate tissue and of the tumour, we selected areas of graded prostate tumours (Gleason score) from cryosectioned preparations and microdissected these cells (20-100 cells). After anionic resin removal of proteins, PCR amplification of TP53 gene exons 5/6 and SSCP analysis, an abnormal SSCP band shift was observed in suspected tumour cells, compared with microdissected stromal cells used as an internal control, while (1) a crude preparation of tissue DNA carrying the tumour did not show any abnormality and (2) immunostaining by a set of monoclonal antibodies against TP53 protein remained negative. Nucleotide sequence analysis of the different bands confirmed the presence of a mutation in the TP53 gene exon 6 position 13,336 in an abnormal band for one specimen, while no mutation was detected in the normal SSCP band. By targeting recognised tumour cells we can find DNA mutations which are undetectable using the standard technique of whole-tissue DNA extraction, particularly in a heterogeneous tumour such as carcinoma of the prostate.


The resultsindicate that all these antibiotics effectively bind copper(ll) at physiological pH. Cyclic voltammetryinvestigations and kinetic studies of H2o2 disproportionation and hydroxyl radicals detection made it possibleto support the mechanism of oxidative reactivity of cupric complexes of aminoglycosides, which involvesCu(1) and Cu(lll) redox states and metal-bound, rather than free radical species. The mechanism of thisprocess appears to be complicated, and may have deleterious side-effects by leaking radical intermediates.The presence of these reactive oxygen species may be responsible for modulating the biological activity ofthese drugs.The majority of aminoglycosidic antibiotics anchor Cu(ll) ions by {NHphe in both the presence and absence of hydrogen peroxideshowed that the complexes with H2o2 are the most efficient oxidants, converting dG to its 8-oxo derivative,generating strand breaks in plasmid DNA and multiple cleavages in tRNAphe. Some of these reactions mayplay a role in aminoglycoside-induced ototoxicity and nephrotoxicity; moreover, they may suggest thatCu(ll)-aminoglycosides are potentially dangerous genotoxic agents.These complexes were also screened for their antibacterial activity and bioassays were engaged to findout the possibility of Cu(ll)-kanamycin A complexes to induce tumor necrosis factor (TNF), interferon (IFN)and interleukin-10 (IL-10) in human peripheral blood leukocytes. The aim of these studies was to comparethe biological action of antibiotic alone and complexed with copper(ll) ions in both neutral and oxidativeenvironment.The interactions of copper(ll) complexes of aminoglycosides with oxidation-susceptible biomolecules:2’-deoxyguanosine, plasmid DNA and yeast tRNA


Concentrations of circulating immune complexes (CIC) were measured serially during chemotherapy of 22 patients with gestational trophoblastic tumours (GTT) and 11 patients with malignant teratoma (MT) by the polyethylene glycol precipitation and CIq solid-phase assays. Results were correlated with tumour response as measured by serum concentrations of human chorionic gonadotrophin (hCG) and alpha-foetoprotein (AFP). CIC concentrations correlated with disease status in the early stages of treatment in 4/22 patients with GTT and 5/11 with MT. CIC assays were less sensitive than hCG and AFP as a monitor of disease, and also less specific, in that 8 patients with GTT and 5 with MT developed raised CIC concentrations during chemotherapy in spite of sustained complete remission. Measurements of CIC concentrations by present methods are neither sufficiently sensitive nor specific to be of clinical value as a tumour marker in GTT and MT, and this casts doubt on their potential value in other malignancies. Attention should be directed to identification of the components of CIC, some of which may be more cancer-specific.


The leukaemic lesions in intact and ovariectomized mice of strain ICRC, induced with 20-methylcholanthrene (20-MCA) in combination with or without hormones were investigated for the presence of mouse leukaemia virus (MuLV) by (i) bioassays and (ii) electron microscopy. The different experimental groups treated with 20-MCA were (i) intact females, (ii) ovariectomized females, (iii) ovariectomized females with pituitary graft, (iv) ovariectomized females with 10 mug oestradiol/day for 30 days and (v) ovariectomized females with 1 mug oestradiol together with 1 mg progesteron/day for 30 days. It was possible to transmit nearly all these experimentally induced leukaemias to syngeneic mice through acellular extracts, compared with very poor transmissibility of spontaneous leukaemias in the ICRC strain, indicating functional activation of viral agents on combined treatment with carcinogen and hormones. Potency of the acellular leukaemic extract from the mice of group (ii) without the ovarian hormones was much weaker than that from mice of the other experimental groups. The leukaemogenic activity of MuLV was enhanced on serial transmission in syngeneic hosts. Leukaemic lesions of ovariectomized mice treated with 20-MCA and oestradiol were also transmissible to the sucklings of allogeneic mice of strain C3H-MTV, C57-BL and Dba-MTV. The cell-free supernatant medium of the cultures of these leukaemic lesions induced leukaemias on back inoculation into syngeneic mice. Electron microscopic studies of lesions induced with carcinogen and oestradiol consistently showed abundant intracytoplasmic type A particles. Numerous intracytoplasmic type A particles as well as some type B particles were found in the leukaemic tissues of ovariectomized females treated with MCA and oestradiol combined with progesterone. Type C particles, characteristic of MuLV were seen in the leukaemic tissues of all other experimental groups. These findings indicate a significant influence of the physiological condition of the host, particularly the hormonal make up, on expression and activity of specific viral agents.


A series of 155 cases of hepatic hydatid disease, occurring in 121 patients, were operated on at the Navaland Veterans Hospital of Athens. Ultrasonography and computerized axial tomography provided thepreoperative diagnosis in 89 and 93 percent of the cases respectively in recent years. Thirty one percentof the cases presented with complications, the commonest of these being infection of the cyst (10percent) and rupture of the cyst into the bile ducts (17 percent). Total cystectomy was performed inthree cases and removal of the endocyst with its content in the remaining 152. The remaining cavity waseither externally drained (57 cases), or filled with omentum (omentoplasty — 95 cases). External fistulaand infection of the residual cavity occurred in 32 and 56 percent after simple drainage and in 4 and 2percent respectively after omentoplasty. Differences are statistically significant (p <  0.001).Hospitalization was also significantly longer after drainage than after omentoplasty (p < 0.01).Obstructive jaundice after intrabiliary rupture of the cyst was more successfully managed afteradditional choledochoduodenostomy than after simple drainage of the common bile duct.Intrapericoneal recurrence of hydatid disease occurred in two cases. The conclusion of the present studyis, that ultrasonography and computerized axial tomography provide an acceptable rate in the diagnosisand that omentoplasty offers a very low complication rate in the management of hydatid cystic disease ofthe liver.


The aim of our research is to evaluate the advantage by the combined use of fiberopticbronchoscopy and laryngeal mask during the performance of percutaneous dilatationaltracheostomy in an intensive care unit.Patients: 16 adult patients who were candidates to middle-long term mechanicalventilation.Environment: Intensive Care Unit of a Community General Hospital.Results: We experienced 3 minor complications (2 minor bleedings and 1 neck emphysema).Difficulties were found in 3 patients with particular anatomical conformation.Conclusion: The combined use of fiberoptic tracheo-bronchoscopy with the laryngealmask permits a better endoscopic visualisation of the operatory field, providing a moresecure and precise procedure.


The aims of this study were: (1) To examine the presence of alphafetoprotein (AFP) bearing tumour cells in PBSC harvests from advanced GCT patients obtained after multiple cycles of induction chemotherapy. (2) To determine whether induction chemotherapy contributed to in vivo purging of the tumour. We evaluated cryopreserved PBSC samples from 5 patients with advanced stage II/III AFP producing GCT. PBSC were separated after the first, second and third cycles of induction chemotherapy. Those samples were analysed using the nested reverse transcription polymerase chain reaction (RT-PCR) method to detect AFP mRNA. Although, in all patients, AFP mRNA was detected in PBSC samples after the first or second cycle of induction chemotherapy, but was not detected in 3 of 4 samples after the third cycle of chemotherapy. Although it is not clear whether tumour cells contaminating PBSC fraction contribute to disease relapse, PBSC harvested after at least 3 cycles of induction chemotherapy might be recommended to avoid such a possibility. © 2001 Cancer Research Campaign


There was an error in the last sentence of the legend to Figure 4.-/- (thin line histogram).The correct text should read: Representative histograms were shown to analyze relative bacterial number in macrophages (MAC) and neutrophils (Neu) between WT (gray-filled histogram) and Rip2


Primary pancreatic lymphoma (PPL) is an extremely rare neoplasm, which may be confused with pancreatic adenocarcinoma. So far only about 150 cases of PPL have been reported.We present our experience of 3 cases of PPL over a 4-year period.All the patients presented with vague abdominal pain of duration ranging from 1½ months to 3 months. Two patients had diagnosis confirmed histologically by CT-guided core biopsy or Fine needle aspiration procedure. We were able to avoid unnecessary laparotomy in 2 patients using preoperative guided Fine needle aspiration Cytology, although the third patient did undergo a Whipple′s procedure as the diagnosis of PPL was not considered during the initial workup.There is no significant difference noted with regard to patient′s age or duration of symptoms between patients with either pancreatic adenocarcinoma or PPL. The differential diagnosis of PPL includes pancreatic adenocarcinoma and secondary involvement of pancreas from extra-nodal lymphoma. Combination of two things is suggestive of Pancreatic lymphoma: (1) Bulky localized tumor in pancreatic head (2) Absence of significant dilatation of main pancreatic duct strengthens a diagnosis of pancreatic lymphoma over adenocarcinoma. Majority of patients can be managed with chemotherapy with much better prognosis compared to patients with pancreatic adenocarcinoma. Larger series of patients are needed to evaluate whether chemotherapy, eventually followed by involved-field radiation therapy, is the treatment of choice for PPL. Gastrointestinal (GI) tract is the most common site of extra-nodal non-Hodgkin′s lymphomas (NHLs), accounting for 15% to 20% of all NHL cases.14 SignifiTo date, approximately 150 cases of PPL have been reported in the English-language medical literature. There isWe present our experience of treating primary pancreatic lymphoma in patients at our hospital. Medical records of patients identified during a retrospective review of prospectively maintained database in the cancer institute of our hospital were retrieved after obtaining informed consent from the concerned patients. A pathologist reviewed slides and blocks to reconfirm the diagnosis. Literature was accessed on the Internet through PubMed search using the string 'primary pancreatic lymphoma.'Over a period of 4 years, we have diagnosed and treated 3 cases of primary pancreatic lymphoma . All theHematoxylin and eosin-stained sections from tumor in all the 3 patients showed monotonous population of small round cells in sheets separated by sparse fibrous septae infiltrating the pancreatic parenchyma . NeoplasTwo patients received chemotherapy  for 4 cycles followed by radiotherapy (45 Gy in 20 fractions for 1 patient and 30 Gy in 18 fractions for the other) using antero-posterior and postero-anterior pair beams. One patient who was diagnosed following Whipple′s procedure received adjuvant chemotherapy with CHOP regimen (6 cycles) alone.All the patients are currently on follow-up  and are so far free of disease recurrence.Diagnostic criteria for PPL include presence of main mass in the pancreas with lymph nodal involvement confined to the peri-pancreatic region with no hepatic or splenic involvement; absence of superficial lymphadenopathy and mediastinal lymph node enlargement; and a normal leukocyte count in peripheral blood.9Literature suggests a strong male predominance .10There iMost common presenting symptom reported in literature is abdominal pain, found in about 83% of cases, followed by (in order of frequency) abdominal mass (58%), weight loss (50%), jaundice (37%), acute pancreatitis 12%), small-bowel obstruction (12%) and diarrhea (12%).131013]


Roquinimex (Roq) is an immunomodulator known to stimulate cellular immune responses. It is currently used for immunotherapy after bone marrow transplantation (BMT). One of the major features of this compound is an enhancement of natural killer (NK) cell activity and numbers. We studied the in vitro effect of Roq on human peripheral blood NK and adherent lymphokine-activated killer cell (ALAK) activities. In cultures supplemented with recombinant interleukin 2 (rIL-2) (1000 U ml-1) and Roq a significant increase in NK and LAK function was observed without a parallel increase in cell numbers. We also examined the generation of NK cells from human bone marrow (BM) immature progenitors, obtained by purging with 4-hydroperoxycyclophosphamide (4HC). NK cell numbers and activity were both increased when cultures with rIL-2 (10 U ml-1) were supplemented with Roq. These results confirm findings obtained in vivo and in vitro in the murine system and suggest that Roq is an active agent on these lymphoid populations. These properties and good tolerability make Roq an attractive tool for immunotherapy.


We discuss the evaluation of the Engaging Older Adult Learners as Health Researchers (ENGOAL), a program designed to educate older adults from underserved and underresourced communities about geriatric health and research methods, enabling them to become Research Partners. Quantitative and qualitative data were collected for evaluation from 21 participants aged 53-79. We used All Aspects of Health Literacy Scale  and an adaptation of Stanford Patient Education Research Center Chronic Disease Self-Efficacy Scale to assess pre-post and follow-up changes, using repeated measures analyses. Results suggest notable increases in self-efficacy, but only small improvements in subjective health literacy. Themes identified through qualitative analysis of interviews with participants included: (1) Finding our Voices (2) Race and Health (3) Faith and Health (4) Communicating with Providers and (5) Sharing and Advocacy. Emerging community leadership of participants are further evidence of confidence gains in our participants.


The evaluation on the characteristic dataset and figures presented here, are related to our latest research data entitled “Fabrication of chitosan based magnetic nanocomposite by click reaction strategy; evaluation of nanometric and Cytotoxic characteristics” [1]. FTIR, Vibrating Sample Magnetometer (VSM) measurements, Xray diffraction (XRD) information and the resulted figures for structural confirmation of the prepared chitosan based nanocomposite are presented in this article. The morphological changes of the Fibroblast, Saos, MCF7 and Hela cell lines after treatment with the mention compound were displayed. The additional adsorption data for the synthesized nanobiocomposite were also demonstrated with graphs. The resulted FT-IR spectra and the corresponding data of the synthesized products were presented in The magnetic nanocomposite was prepared using chitosan. To prepare functionalized chitosan, the chitosan was azidated using chloroacetyl chloride and sodium azide. Then click reaction which has been incorporated in our recent studies ,3 and als) of 25.4 (emu/g) has super paramagnetic characteristics. Moreover; the corresponding data were presented in Magnetization experiments of the prepared magnetic nanocomposite (MNC) were obtained using VSM technique at room temperature. As can be seen in 3O4 nanoparticles  theory were performed. The isotherm plots were used to calculate the specific surface area and the average pore diameter of the chitosan/magnetic nanocomposite and the difference between adsorption and desorption steps.For evaluating the cell cytotoxicity of the prepared sample (MNC) according to the literature , some kn


Qualitative research on positive coping approaches actually used by caregivers can inform interventions that can be feasibly implemented. Absent from previous qualitative research is how caregivers respond to strain in the relationship, specifically. Eight focus groups were conducted with a purposeful sample of racially and ethnically diverse family caregivers in Los Angeles (n=75). An additional 8 in-depth follow-up interviews were conducted. Content analysis was used to understand the mechanisms employed by caregivers to cope with strain and tension in the caregiving relationship. Preliminary results revealed twenty-two individual themes, which were subsequently grouped into four main superordinate themes: 1) Self-care; 2) Adaptation of behaviors and feelings; 3) Seeking and utilizing assistance and respite; and 4) Education and support groups. This work can help inform the design of programs to support caregivers and prevent potentially harmful behaviors, through understanding the experiences of caregivers in their own words.


Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer (BC). It is so named because it lacks expression of estrogen/progesterone receptors and does not overexpress the human epidermal growth factor receptor 2 (HER2). While only 15–20% of all BC cases are classified as TNBCs, they account for over 50% of BC mortality. TNBCs include the BRCA1/2- defective BCs and largely overlap with the basal-like BC group. As of today, no targeted therapies are available for these BC subtypes and despite advances in cytotoxic therapies they still carry a grim prognosis, with relapses typically occurring as soon as 3 years after treatment [TNBCs trace their origin to the genomic instability (GI) associated with defects in DNA repair (notably homology-directed repair (HDR)) and thus, compared to other BCs show increased rates of pathologic complete response following chemotherapy . Yet, deDNM2 was associated with lower relapse-free survival and very short times (3–5 years) to relapse after chemotherapy in TNBC. Below we provide a mechanism that explains how increased DNM2 expression may shape resistance in later stages of the disease and why it is so uniquely important for the outcome in TNBC.We have recently shown that the efficiency of HDR depends on dynamin 2 (DNM2), a large GTPase best known for its role in intracellular molecular trafficking [RAD51 might similarly reverse the HDR defects induced by DNM2 deficiency. However, although we successfully rescued the DNM2 inhibition-induced HDR phenotypes in all other cells, this was not the case for cells derived from advanced TNBCs, which remained strikingly dependent on DNM2 function for their survival after treatment with chemotherapy. We concluded that in the absence of BRCA and other proteins that control recruitment of RAD51 to the sites of DNA damage and/or stalled replication forks, RAD51 trafficking to the nucleus and subsequent cell resistance to chemotherapy was largely dependent on DNM2. This reasoning is supported by observations of aberrant cytoplasmic-to-nuclear ratios of RAD51 in late-stage TNBCs, suggesting the importance of RAD51 trafficking between nucleus and cytoplasm for the aggressiveness of TNBC [High genomic instability (GI) is a characteristic feature of TNBCs. One mechanism often employed by TNBCs to counteract GI is overexpression of RAD51, a protein central to HDR . As a DN of TNBC .Recent single-cell sequencing of longitudinal TNBC samples  has showIn summary, increased DNM2 and associated intra-cell trafficking explain how DNA repair-deficient cells could acquire both resistance to chemotherapy and mobility, thus providing one possible solution to the triple-negative paradox of BC (Figure


Here, we investigate cross-seeding of full-length human Tau (FL Tau) with fibrils assembled from K18 and K18ΔK280 fragments of Tau in the presence of poly-L-glutamate (poly-Glu) as an enhancer of Tau aggregation. To study cross-seeding between Tau polypeptides and the role of the conformational memory effect in induction of Tau amyloid polymorphism, kinetic assays, transmission electron microscopy, infrared spectroscopy and limited proteolysis have been employed. The fastest fibrillization was observed for FL Tau monomers seeded with preformed K18 amyloid yielding daughter fibrils with unique trypsin digestion patterns. Morphological features of daughter FL Tau fibrils induced by K18 and K18ΔK280 seeds were reminiscent of the mother fibrils , respectively) but disappeared in the following generations which became similar to unpaired FL Tau amyloid fibrils formed de novo. The structural evolution observed in our study was accompanied by disappearance of the unique proteolysis profile originated from K18. Our findings may have implications for understanding molecular mechanisms of the emergence and stability of Tau amyloid strains.Amyloid aggregates of Tau protein have been implicated in etiology of many neurodegenerative disorders including Alzheimer's disease (AD). When amyloid growth is induced by seeding with preformed fibrils assembled from the same protein, structural characteristics of the seed are usually imprinted in daughter generations of fibrils. This so-called conformational memory effect may be compromised when the seeding involves proteins with non-identical sequences leading to the emergence of distinct structural variants of fibrils (amyloid ‘ In A and B, lane 1 shows molecular weight marker. Note different pattern of bands in the range of 15 to 20 kDa in lane 3.(TIF)Click here for additional data file.


The study examined attitudes towards Advance Care Planning (ACP) as a potential mediator in the association between acculturation and willingness to use nursing home in Korean American older adults (aged 60 and older). Data were driven from a cross-sectional study of 235 community-dwelling Korean American older adults (aged 60 and older) in 2013. Multivariate regression models of willingness to use nursing home were entered in the following order: (1) demographics, (2) health, (3) acculturation, and (4) attitudes towards advance care planning. The mediation effect of attitudes towards ACP in the relationship between acculturation and willingness to use nursing home was separately examined using the bootstrapping method. Higher acculturation was associated with positive attitudes towards ACP and more likelihood of using nursing home. The proposed mediation model was fully supported: positive attitudes towards ACP served as an intervening step between acculturation and willingness to use nursing home. The mediating role of attitudes towards ACP yields implications for developing culturally sensitive advance care planning education program targeting older individuals.


A 39-year-old man presented with a productive cough, a history of chronic dyspnea and recurrent lower respiratory tract infections. Physical examination and laboratory tests were unremarkable. A chest X-ray (A) showed an enlarged tracheal diameter (white arrows), with multiple cysts in the lower lobes (red arrows). A computed tomography scan of the chest (B) showed the dilatation of both the trachea and the two main bronchi (white arrows) with multiple diverticulae (blue arrows). It also revealed bilateral cystic bronchiectasis involving both lower lung zones (red arrows). A possibility of Mounier-Kuhn syndrome (MKS) was considered. MKS is a rare condition characterized by recurrent lower respiratory tract infections and tracheobronchial dilation that is due to atrophy of the muscular and elastic tissues in the trachea and main bronchial wall. The patient underwent bronchoscopy which demonstrated tracheal dilation with diverticulae and enlargement of both main bronchi confirming the diagnosis of MKS.


The management of multiple chronic conditions (MCC) in older adults living in the community is complex. Little is known about the experiences of interdisciplinary primary care and home providers who care for this vulnerable group. The aim of this study was to explore the experiences of healthcare providers in managing the care of community-living older adults with MCC and to highlight their recommendations for improving care delivery for this group. A qualitative interpretive description design was used. A total of 42 healthcare providers from two provinces in Canada participated in semi-structured interviews. Participants represented diverse disciplines  and settings . Thematic analysis was used to analyze interview data. The experiences of healthcare providers managing care for older adults with MCC were organized into six major themes: (1) managing complexity associated with MCC, (2) implementing person-centred care, (3), involving and supporting family caregivers, (4) using a team approach for holistic care delivery, (5) encountering rewards and challenges in caring for older adults with MCC, and (6) recommending ways to address the challenges of the healthcare system. Healthcare providers highlighted the need for a more comprehensive integrated system of care to improve care management for older adults with MCC and their family caregivers. Specifically, they suggested increased care coordination, more comprehensive primary care visits with an interprofessional team, and increased home care support.


Recent data from an individual patient data (IPD) meta-analysis of 17 randomized control trials including >100,000 women living in low- and middle-income countries found that multiple micronutrient supplementation (MMS) in pregnancy reduced the risk of low birth weight, preterm birth, and being born small for gestational age. Further, MMS reduced the risk of neonatal and infant mortality for females, and there was no evidence of increased risk among the 26 subgroups examined. The 2016 WHO antenatal care guidelines, which were released before the IPD meta-analysis, did not universally recommend MMS, noting: “There is some evidence of additional benefit … but there is also some evidence of risk.” The guidelines suggest that MMS may increase the risk of neonatal mortality based on an exploratory subgroup analysis of 6 randomized trials. However, we identified several issues with this subgroup analysis. In this report we correct and update the subgroup analysis and show that there is no evidence that MMS increases the risk of neonatal mortality. There is growing scientific consensus that MMS containing iron and folic acid (IFA) is superior to IFA alone. The WHO guidelines currently state that “policy-makers in populations with a high prevalence of nutritional deficiencies might consider the benefits of MMN [multiple micronutrient] supplements on maternal health to outweigh the disadvantages, and may choose to give MMN supplements that include iron and folic acid.” This equivocal guidance has created confusion about the best course of action for public health programs in low- and middle-income countries. Given the new evidence, WHO should review their statements regarding the potential neonatal mortality risks and re-evaluate the overall potential benefits of implementing MMS as a public health program. Micronutrient deficiencies in pregnancy are common and are associated with adverse birth outcomes . PrenataThe 2016 WHO antenatal care guidelines do not universally recommend MMS, noting: “There is some evidence of additional benefit of MMN [multiple micronutrient] supplements containing 13–15 different micronutrients (including iron and folic acid) over iron and folic acid supplements alone, but there is also some evidence of risk” (1) corrected the Bhutta et al. estimate, which is consistent with the updated 2017 Cochrane Review (2) included the omitted MINIMat study (3) added 2 recently published trials (lipid-based supplements arms excluded) (4) included 2 trials that used a 60 mg Fe/d control but were excluded due to using a 250 µg/d folic acid dose  addexcluded) , 7; and cid dose , 9. The cid dose  and was cid dose . The Fawcid dose  trial wacid dose  neonatalcid dose –16, and cid dose .and the effect of additional micronutrients. The lower dose of iron in the MMS (30 mg/d) as compared with the control arm (60 mg/d) may explain negative effect estimates seen in some trials. The 2017 Cochrane Review examined the effect of MMS on perinatal mortality stratified by iron dose in the MMS and control arms  with an IFA control arm containing high-dose iron (60 mg/d). These trial designs simultaneously address 2 questions about the effect of a lower dose of iron rol arms . Among trol arms . We argudecrease the risk of infant mortality for infants born to women without access to a skilled birth attendant (∼18% risk reduction)  . The IPDOur update of the WHO subgroup analysis showed no increased risk of neonatal mortality and the results are consistent with the findings of the updated 2017 Cochrane Review and our recent IPD meta-analysis which found that MMS did not increase the risk of neonatal mortality overall or in any of 26 subgroups of pregnant women and newborns . In fact


SLE and Serum Complement: Causative, Concomitant or Coincidental?2017, 11: 113-122The Open Rheumatology Journal, The revised last paragraph of conclusion in abstract is mentioned below:It is clinically important to find novel ways to assess disease activity in SLE. Increased serum levels of cell-bound complement activation products may more accurately reflect disease activity than conventional serum C3 and C4 monitoring.The original last paragraph of conclusion provided was:It is clinically important to find novel ways to assess disease activity in SLE. Reduced serum levels of cell-bound complement activation products may more accurately reflect disease activity than conventional serum C3 and C4 monitoring.The revised last paragraph of conclusion is mentioned below:With recent studies demonstrating that increased levels of serum cell-bound complement activation products may more accurately reflect disease activity than conventional complement C3 and C4 monitoring, we believe that this is an important area for future SLE research and look forward to further studies on research in the complement in SLE.The original last paragraph of conclusion provided was:With recent studies demonstrating that reduced levels of serum cell-bound complement activation products may more accurately reflect disease activity than conventional complement C3 and C4 monitoring, we believe that this is an important area for future SLE research and look forward to further studies on research in the complement in SLE.


Parkinson’s disease (PD), a common neurodegenerative disorder affects approximately 1% of the population over 65. PD is a late-onset progressive motor disease characterized by tremor, rigidity (stiffness), and bradykinesia (slowness of movement). The hallmark of PD is the selective death of dopamine-containing neurons in the substantia nigra pars compacta which send their projections to the striatum and the presence of cytoplasmic aggregates called Lewy bodies [1-2]. Most cases of PD are sporadic but rare cases are familial, with earlier onset. The underlying mechanisms and causes of PD still remain unclear.


Macromolecular complexes are essential to intracellular signal transduction by creating signaling niches and enabling a chain of reactions that transmit external signals into various cellular responses. Analysis of SMYD3 interactome indicates this protein lysine methyltransferase might be involved in calcium dependent signaling pathways through forming complexes with the phospholipase PLCB3, calcium/calmodulin dependent kinase CAMK2B, or calcineurin inhibitor RCAN3. SMYD3 is well-known as a histone H3K4 methyltransferase involved in epigenetic transcriptional regulation; however, any roles SMYD3 may play in signaling transduction remain unknown. KEGG pathway enrichment analysis reveals the SMYD3 interacting proteins are overrepresented in several signaling pathways such as estrogen signaling pathway, NOD-like receptor signaling pathway, and WNT signaling pathway. Sequence motif scanning reveals a significant enrichment of PXLXP motif in SMYD3 interacting proteins. The MYND domain of SMYD3 is known to bind to the PXLXP motif. The enrichment of the PXLXP motif suggests that the MYND domain is likely to be a key interaction module that mediates formation of some SMYD3 complexes. The presence of the PXLXP motifs in PLCB3 and CAMK2B indicates the potential role of the MYND domain in mediating complex formation in signaling. The structural basis of SMYD3 MYND domain-mediated interactions is unknown. The only available MYND-peptide complex structure suggests the MYND domain-mediated interaction is likely transient and dynamic. The transient nature will make this domain well-suited to mediate signaling transduction processes where it may allow rapid responses to cellular perturbations and changes in environment. The N-tThe structure of SMYD3 in complex with a MAP3K2 peptide provided the structural basis for SMYD3-substrate complex formation . The weak interaction suggests the MYND-mediated interactions are transient in nature. Transient interactions usually show a fast bound-unbound equilibrium and can readily switch between different binding partners (Perkins et al., MYND domain-based interactions show relatively low binding affinities with KThe structural basis of SMYD MYND domains-mediated interactions is unknown. Based on the structural superposition , the pepIn contrast to binding specificity, MYND domains also show promiscuous binding. Many MYND domains are capable of binding to multiple targets, and single peptide motifs can be recognized by different MYND domains. The MYND domain of BS69 is able to bind to MGA, E1A, and EBNA2 proteins (Ansieau and Leutz, Motif scanning using ScanProsite (de Castro et al., The significant enrichment of PXLXP motif in SMYD3 interacting proteins suggests that the MYND domain is likely to be a key interaction module that mediates the formation of some SMYD3 complexes. The presence of PXLXP motifs in PLCB3 and CAMK2B indicates the potential role of the MYND domain in mediating complex formation in signaling including the aforementioned estrogen signaling pathway, GnRH signaling pathway, NOD-like receptor signaling pathway, and WNT signaling pathway. In PLCB3 there are two PXLXP motifs, PPLRP at residues 265–269 and PSLEP at residues 530–534 . The for2+ concentrations; as intracellular Ca2+ concentrations rise, the holoenzyme is released from the cytoskeleton and enters the cytosol (Swulius and Waxham, CAMK2B has only one PXLXP motif, PCLSP at residues 476–480. This motif is located at the variable segment of CAMK2B . The varPseudomonas aeruginosa-induced intracellular calcium signaling and regulates the activation of PKC and the nuclear transcription factor NF-κB in human bronchial epithelial cells (Bezzerri et al., Analysis of SMYD3 interactome reveals SMYD3 might be involved in signal transduction pathways in addition to epigenetic gene regulation. Most enriched KEGG pathways are associated with the signaling processes in cancer. Among them, the WNT signaling has been shown to affect the maintenance of stemness and metastasis of cancer cells (Zhan et al., Caenorhabditis elegans orthologs, Bra-1 and Bra-2, have been shown to participate in BMP and TGFβ signaling pathways (Kurozumi et al., The MYND domain might be a key complex-forming module in SMYD3 complexes. The PXLXP motif to which the MYND domain may bind is significantly enriched in SMYD3 interacting proteins. However, there is no significant difference in the enrichment between the interactors involved in signaling transduction pathways and epigenetic transcriptional regulation suggesting that the MYND domain does not have any particular binding preferences in either of these processes. No MYND domain has been directly linked to signaling, but some MYND domain-containing proteins such as BRAM1 as well as its SMYD3 interacting proteins were retrieved from four protein-protein interaction databases including GPS-Prot (Fahey et al., Pathway enrichment analysis of SMYD3 interactors was performed using STRING web-based analyzer (Szklarczyk et al., The presence of PXLXP motif in SMYD3 interacting proteins was scanned using ScanProsite (de Castro et al., All datasets generated for this study are included in the article/supplementary material.ZY and YZ contributed conception and design of the study, performed the database and statistical analyses, and prepared the figures. ZY wrote the first draft of the manuscript. All authors contributed to manuscript revision, read, and approved the submitted version.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.


Candida spp., the rate of acute kidney injury (AKI) defined by a creatinine above 3.5 mg/dl or the need of any form of renal replacement therapy (RRT) was around 20% in the bacteria group versus above 40% in the candida group, while having similar Acute Physiology and Chronic Health Evaluation (APACHE) II scores [d-glucan (BDG) could be a very good candidate associated to PCT. Indeed, BDG could be even a better candidate as its molecular weight ranges from several hundred thousand to 10 million daltons and does not pass through any membrane [Candida albicans that increases serum BDG without candidemia [In their recent systematic review, Cortegiani et al. found that serum procalcitonin (PCT) concentrations were higher in patients with bacteremia as compared to candidemia . QualityAndrea Cortegiani, Mariachiara Ippolito and Antonino GiarratanoCandida spp. receiving CRRT and blood purification techniques should be addressed in future studies. Another interesting aspect would be the impact of CRRT in the trend of PCT values in these patients, since almost all the included studies reported the values at the beginning of the diagnostic process [We would like to thank Dr. Honore et al. for discussing the potential role of acute kidney injury (AKI) and continuous renal replacement therapy (CRRT) as potential confounders on the different values of procalcitonin (PCT) in patients with bacteremia versus candidemia in the intensive care unit (ICU) in our systematic review . We eval process . However process –9. These


The efficiency of organotin(IV) complexes as photostabilizers was determined by monitoring the changes in the weight, growth of specific functional groups , viscosity, average molecular weight, chain scission and degree of deterioration of the polymeric films upon irradiation. The results obtained indicated that organotin(IV) complexes stabilized poly(vinyl chloride) and the dimethyltin(IV) complex was the most efficient additive. The surface morphologies of poly(vinyl chloride) films containing organotin(IV) complexes were examined using an atomic force microscope and scanning electron microscopy. These showed that the surface of polymeric films containing organotin(IV) complexes were smoother and less rough, compared to the surface of the blank films. Some mechanisms that explained the role of organotin(IV) complexes in poly(vinyl chloride) photostabilization process were proposed.Three organotin(IV) complexes containing ciprofloxacin as a ligand (Ph The market for poly(vinyl chloride) (PVC) has grown significantly over the years due low production cost and its large versatility ,2,3. HowA UV light of a wavelength longer than 190–220 nm is not expected to be absorbed by PVC since it only contains single bonds  . CommercAs a continuation of our research in the field of polymers ,38,39,403SnL, Me2SnL2 and Bu2SnL2; 2SnL2 and Bu2SnL2) and triorganotin (Ph3SnL) were used. Also, the substituents on the Sn were varied as small (Me) and bulky (Bu and Ph) groups.Three organotin-ciprofloxacin complexes  complex is the least sterically hindered among the additives used and, therefore, it acts as a better primary photostabilizer compared to the others. Triphenyltin complex shows the least photostabilization effect presumably due to the presence of bulky phenyl groups.Photodegradation of PVC produces low molecular weight fragments as well as hydrochloride (HCl) that lead to a weight loss . The wei−1), C=O (1722 cm–1) and C=C (1602 cm−1) , can be used to evaluate the changes in PVC average molecular weight [η]=The PVC molecular weight was calculated from the intrinsic viscosities using Equation (6).Three organotin(IV) complexes containing ciprofloxacin as a ligand have been used as photostabilizers to inhibit the photodegradation of PVC films, upon irradiation with ultraviolet light for a long period. Various methods such as the growth of certain functional groups in the infrared spectra, percentage of weight loss, variation in molecular weight, chain scission and degree of deterioration were used to determine the efficiency of organotin(IV) complexes. In addition, the atomic force and scanning electron microscopy were used to assess the surface morphology of poly(vinyl chloride) films. The organotin(IV) complexes used act as efficient photostabilizers and reduce the photodegradation rate of poly(vinyl chloride). It is believed that organotin(IV) complexes act as hydrogen chloride scavengers, peroxide decomposers, free radical scavengers and UV absorbers. For future research using organotin complexes as PVC photostabilizers, the possible leakage of tin should be assessed in order to allow the long-term use of such additives.


The discovery of the involvement of defined molecular pathway(s) in tumor development and therapeutic targeting of such pathway(s) has revolutionised cancer treatment. More than 50 targeted therapeutic agents have been approved by the Food and Drug Administration (FDA) in the last two decades . Unlike ALK G1202R mutation. It is also highly permeable through the blood brain barrier and hence effective in targeting central nervous system metastasis. Recently published results of the global phase II study has shown superior objective response rate (ORR) and intracranial ORR in comparison to the first and second generation ALK TKIs [V600E/K melanoma patients that relapse on monotherapy [Anaplastic Lymphoma Kinase (ALK) exemplifies the use of targeted therapeutic agents in combating cancer, as well as the challenges that arise in terms of resistance mechanisms against such agents in clinical settings . A numbeALK TKIs . As the ALK TKIs . OrganicALK TKIs , 9. One otherapy . However


In the third (HSP90(Cyt)) and fourth (HSP60(Mit)) panels of Both the HTML and PDF versions of the article have been corrected. This error appears only in print and in PDF versions downloaded on or before March 28, 2019.


Although significant animal suffering caused by preventable diseases is frequently seen in developing countries, reports of this are scarce. This report describes avoidable animal suffering owing to a suspected canine distemper (CD) outbreak in unvaccinated dogs owned by low-income families in Mozambique that killed approximately 200 animals. Affected dogs exhibited clinical signs, and gross and microscopic lesions compatible with CD. Immunohistochemical staining confirmed the presence of canine distemper virus (CDV) in the kidney of one dog from the cohort. This brief communication again illustrates that large outbreaks of CDV in unvaccinated dogs occur and that large-scale avoidable suffering and threats to the health of dogs and wild canines continue. Mass vaccination supported by government and non-government organisations is recommended. Morbillivirus within the subfamily Paramyxovirinae.Canine distemper (CD) is a highly contagious, acute or subacute systemic viral disease that has a high mortality rate in dogs and wild canines. It is caused by canine distemper virus (CDV), which belongs to the genus et al. et al. et al. et al. et al. et al. et al. Vaccination remains the principal strategy for protection, and once clinical signs develop, treatment is limited to supportive care. Even in settings where a diagnosis is rapidly reached, a high standard of care is available and high levels of vaccination are achieved, CD outbreaks continue to occur amongst domestic dogs .The most obvious gross post-mortal lesions were cachexia (3/3), mucopurulent exudate in the trachea and bronchi (2/3), focal pneumonia (3/3), cyanotic heart (2/3), splenic atrophy with increased consistency (2/3), pale kidneys (3/3) and contracted bladders with multifocal haemorrhages in the mucosa (2/3).Histopathological examination of the lung, bronchi and bronchioli showed extensive areas of severe purulent inflammation (2/3). Splenic and lymph node tissues appeared moderately active to highly active (2/3). In the grey matter of the cerebellum, multifocal blood vessels showed notable leukostasis, predominantly with lymphocytes and a few macrophages, and there were also mononuclear infiltrates visible in the perivascular spaces in these areas. There was mild gliosis in the tissue. A few blood vessels in the meninges also showed perivascular cuffing with mononuclear leukocytes, predominately lymphocytes (2/3). A few multifocal aggregates of neutrophils were visible in the parenchyma of the liver, and a mild degree of leukostasis was evident in the sinusoids. The portal regions showed the presence of leukocytes (3/3). Focally extensive severe interstitial inflammation of a chronic lymphoplasmocytic appearance was seen in the prostate (3/3).The history of the disease, the clinical signs and gross and histopathological presentations raised the probability of the presence of a CDV infection. To support this putative diagnosis, immunohistochemistry was performed, which confirmed the disease through positive immunoreactivity to CDV antigens in one of these three cases.et al. et al. The incidence of CDV infections worldwide has decreased thanks to the introduction of the highly protective CDV-modified live virus (MLV) vaccines more than 60 years ago (Martella This report illustrates the highly contagious nature of the disease and the threat to animal welfare posed by non-vaccination. Valuable wildlife would also potentially be threatened by this disease in dogs.


This methodological paper discusses the process of co-creating a documentary film with seven formerly homeless older adults, highlighting some of the tensions carrying out community-based participatory research (CBPR). This paper is part of a larger study that explored ‘finding home’ through a series of individual and group audio and video-recorded interviews  with seven adults (aged 50+) with diverse homeless histories. In addition to the main findings, participants shared their experience of filmmaking and CBPR. Findings revealed four main tensions: 1) openness of sharing stories versus privacy and anonymity; 2) balancing participation/engagement and over-burdening; 3) negotiating interpersonal conflict and community building; and 4) ethical issues surrounding copyright and ownership of the film. Ultimately, we advocate for more CBPR film projects, as they not only provide a rich contextualized window into people’s everyday lives but serve to advance the voices of marginalized populations beyond traditional academic circles.


OBJECTIVES/SPECIFIC AIMS: Acculturation and discrimination are associated with negative health outcomes including functional disability. The effect of these on functional disability in older African immigrants in the United States is not well understood. The purpose of this study is to describe and examine the experiences of acculturation, racial discrimination and functional disability in older African immigrants. METHODS/STUDY POPULATION: This study will use cross-sectional quantitative and qualitative mixed methods design. We plan to recruit 150 older (>55 years) participants through the African Immigrant Health Study and community-based organizations serving African immigrants. We will use the PROMIS physical function measure to assess functional disability, the Everyday Discrimination scale and the Psychological Acculturation scale will be used to measure discrimination and acculturation respectively. Higher scores indicate greater severity. RESULTS/ANTICIPATED RESULTS: We have recruited 12 participants so far. The mean age is 57 years and mean length of stay in the United States is 23 years. Mean disability score is 6.5 (range 1–38). Mean discrimination is 8.2 (range 4–15). The prevalent acculturation strategy of these participants (7) is marginalization (neither identified with the American nor African cultures). DISCUSSION/SIGNIFICANCE OF IMPACT: Preliminary results indicate pervasive discrimination and marginalization of study participants. Exploring these experiences can inform preventive strategies of coping and health behaviors that can decrease the negative effects of discrimination, acculturation and functional disability in African immigrants.


Roughly one-third of dissolutions among married persons aged 50 and older occur through divorce rather than widowhood, reflecting the rising gray divorce rate and lengthening life expectancies. We use data from the 1980 Vital Statistics and the 2017 American Community Survey (ACS) to estimate the divorce and widowhood rates among married individuals (aged 50+) in 1980 and 2017 to track how much the widowhood rate has declined and the divorce rate has risen. In 1980, women’s widowhood rates exceeded their divorce rates at all ages. For men, the rate of divorce outpaced the rate of widowhood through ages 50-54. By 2017, divorce rates were higher for women through ages 55-59 and for men through ages 60-64, coinciding with the growth in gray divorce. We also examine subgroup variation in the 2017 patterns and the sociodemographic correlates of having experienced divorce versus widowhood during the past year using the ACS data.


This study aims to understand the roles that psychosocial factors play on the longitudinal associations between sensory (including hearing and vision) loss and cognitive decline. Specifically, we hypothesized that (1) loneliness mediates the associations between sensory loss and cognitive decline; and (2) social support moderates the associations between sensory loss and cognitive decline. We used longitudinal parallel process (LPP) modeling with data from the Health and Retirement Study (HRS) and the Aging, Demographics, and Memory study (ADAMS). Age variable centered at its mean age of 82. In the most parsimonious model, loneliness fully mediated the associations between vision loss and the average cognitive status at age 82 (p < .05). Social support moderated the associations between vision loss and the average cognitive status at age 82 . No moderation or mediation effect was found for the psychosocial factors on the associations between hearing loss and cognition.


The last poliomyelitis cases reported in Papua New Guinea occurred in 1996. Papua New Guinea is one of 37 countries (or areas) of the World Health Organization Western Pacific Region that were certified free of indigenous wild poliovirus in 2000. On June 22, 2018, the National Department of Health confirmed an outbreak of poliomyelitis caused by circulating vaccine-derived poliovirus type 1 (cVDPV1) following isolation of genetically linked virus from a patient with paralysis and nonhousehold community contacts. The index patient was a boy aged 6 years from Lae, Morobe Province, with onset of paralysis on April 25 and history of having received 2 doses of Sabin oral poliovirus vaccine (OPV).cVDPVs can emerge in underimmunized populations when Sabin vaccine poliovirus is extensively transmitted person-to-person and reverts to neurovirulence  (Round 1 and Round 2)  with bivPapua New Guinea was at risk for delayed detection of poliovirus because of an insufficient surveillance system for acute flaccid paralysis (AFP) and delays in seeking health care, often because of geographic inaccessibility. After the initial cVDPV1 case was detected, active AFP case-finding at health facilities was intensified. The annual national nonpolio AFP rate, a key indicator of surveillance sensitivity (A December outbreak response assessment concluded that cVDPV1 transmission likely continues, given the dates of isolation of cVDPV1 from environmental surveillance and the most recent confirmed case. Additional SIAs are planned in Papua New Guinea in 2019. Because of the outbreak, on July 12, 2018, CDC issued a Level 2 Travel Health Notice recommending that all travelers to Papua New Guinea be fully vaccinated against polio. Before traveling to Papua New Guinea, adults who completed their routine polio vaccine series as children are advised to receive a single, lifetime adult booster of polio vaccine.


Following publication of the original article , the autOriginally, the sentence was stated as:(ii) exhibited no evidence of prostate cancer as determined by blood test (PSA < 4 mg/dL)The correct sentence is:(ii) exhibited no evidence of prostate cancer as determined by blood test (PSA < 4 ng/ml)


Rhinusa and Gymnetron, two closely related groups of weevils that feed and develop inside plant tissues of hosts within the families Scrophulariaceae and Plantaginaceae. We found strong evidence for phylogenetic conservatism of host use at the plant family level, most likely due to substantial differences in the chemical composition of hosts, reducing the probability of shifts between host families. In contrast, the use of different plant organs represents a more labile ecological trait and ecological niche expansion that allows a finer partitioning of resources. Rhinusa and Gymnetron weevils initially specialized on plants within Scrophulariaceae and then shifted to the closely related Plantaginaceae; likewise, a gall inducing behavior evolved from non-galler weevils, possibly in response to resource competition, as galls facilitate larval development by providing enhanced nutrition and a favorable microhabitat. Results from trait-dependent diversification analyses suggest that both use of hosts within Plantaginaceae and parasitism on fruits and seed capsules are associated with enhanced diversification of Rhinusa and Gymnetron via low extinction rates. Our study provides quantitative evidence and insights on the ecological factors that can promote diversification in phytophagous insects that feed and develop inside plant tissues.Weevils (Coleoptera: Curculionoidea) represent one of the most diverse groups of organisms on Earth; interactions with their host plants have been recognized to play a central role in their remarkable diversity, yet the exact mechanisms and factors still remain poorly understood. Using phylogenetic comparative analyses, here we investigate the evolution of host use and its possible role in diversification processes of  Comprising approximately 5,800 genera and more than 60,000 described species , weevilsRhinusa and Gymnetron (Curculionidae: Curculioninae), endophagous parasitic weevils whose larvae feed and develop within tissues of plant species in the families Scrophulariaceae and Plantaginaceae. The genus Rhinusa comprises approximately 40 species with a Palearctic distribution  as host plants  assess phylogenetic conservatism in host use by plant family and in different modes of parasitic behavior; (2) estimate ancestral states of traits associated with host plant use; and (3) assess possible effects of these traits on speciation and extinction rates within the Rhinusa/Gymnetron species complex. These analyses should provide valuable insights into the evolution of different traits of host plant use and their role in diversification processes of phytophagous insects that feed and develop inside plant tissues.Here, we use a previously inferred phylogeny from mitochondrial and nuclear DNA sequence data  in conjuRhinusa representing approximately 80% of recognized species, with representatives from the three main taxonomic groups proposed by Gymnetron from the Palearctic and Afrotropical regions (approximately 40% of recognized species). Additionally, since some of the analyses performed in this study require ultrametric trees, we performed a Bayesian analysis for the same dataset of concatenated sequences using BEAST 2.1.3 ST 2.1.3 .We used Mesquite 3.1  to assesUsing the same host plant use categorization as for phylogenetic conservatism analyses, ancestral states were reconstructed across the random sample of 1,000 post burn-in trees from the analysis performed with MrBayes. We used the Bayesian approach reversible jump MCMC (rjMCMC), implemented in BayesTraits 2.0 ; the advFor comparison with the rjMCMC approach and to account for differential speciation, extinction and character transition rates in the reconstruction of ancestral states, we additionally performed ancestral state reconstructions using maximum likelihood under binary and multi-state speciation and extinction models (BiSSE/MuSSE)  in the RRhinusa/Gymnetron species complex. We independently used the model to assess the effect of host plant family use (Scrophulariaceae or Plantaginaceae) and the mode of parasitism as gall or non-gall inducer. The model estimates six parameters; speciation (λ) and extinction (μ) rates for states 0 and 1  and transition rates from state 0 to 1 and vice versa (q01 and q10). Similarly, a multi-state speciation and extinction (MuSSE) model was employed to assess the effect of different modes of parasitism on either roots, stems or fruit/seed capsules. The MuSSE model is an extension of the BiSSE model to discrete traits with more than two states or combinations of binary traits analyzed simultaneously . Eight parsimony steps were observed against 14 expected steps at the lower confidence limit under 10,000 character randomizations of the null model. When host plant use is categorized according to the plant parts being parasitized and as gall inducer vs. non gall inducer, the distribution of characters over the Bayesian consensus tree is marginally significant at the 0.05 confidence level, that is, in both cases the difference between the expected (null model) and observed number of parsimony steps is only one. A similar trend was observed for the results from the estimation of Pagel’s lambda parameter, which indicate a strong phylogenetic signal for host use by plant family (λ = 1) but less phylogenetic signal for both host use as either gall or non-gall inducer and parasitic mode on different plant structures , providing enhanced nutrition and a favorable microhabitat which facilitates larval development. The use of restricted sets of host plants is phylogenetically conserved, most likely because of substantial differences in the chemical composition of their hosts, thus reducing the probability of host shifts. In contrast, the utilization of different plant organs represents a more labile ecological trait that allows a finer partitioning of resources; this ecological niche expansion is associated with enhanced diversification rates in weevils exhibiting a parasitic behavior on fruits and seed capsules. Our results show that ecological factors such as host plant use and specialized endoparasitic habits can promote diversification via low extinction rates in phytophagous insects that require plant tissues for the completion of their reproductive cycle.10.7717/peerj.6625/supp-1Supplemental Information 1Extinction rates associated with (A) use of either Scrophulariaceae or Plantaginaceae as host plant families and (B) use of different plant organs: roots, stems or fruits and seed capsules.Click here for additional data file.10.7717/peerj.6625/supp-2Supplemental Information 2Extinction rates associated with use of Plantaginaceae taxa as hosts (A) and use of fruits and seed capsules (B) estimated with MCMC analyses (BiSSE and MuSSE models) from pruned simulated trees. The red line indicates the true parameter value used to simulate the tree.Click here for additional data file.


We deeply appreciate the great contributions of many physicians in the registry of esophageal cancer cases. The Comprehensive Registry of Esophageal Cancer in Japan, 2012, was published here, despite some delay. The registry complies with the Act for the Protection of Personal Information. The encryption with a HASH function is used for anonymity in an unlinkable fashion.We briefly summarized the Comprehensive Registry of Esophageal Cancer in Japan, 2012. Japanese Classification of Esophageal Cancer 10th [We hope that this Comprehensive Registry of Esophageal Cancer in Japan for 2012 will help to improve all aspects of the diagnosis and treatment of esophageal cancer in Japan.I.Clinical factors of esophageal cancer patients treated in 2012Institution-registered cases in 2012Patient backgroundTableAge and genderTablePrimary treatmentTableTumor locationTableHistologic types of biopsy specimensTableDepth of tumor invasion, cT (UICC TNM 7th)TableLymph node metastasis, cN (UICC TNM 7th)TableDistant metastasis, cM (UICC TNM 7th)TableClinical stage (UICC TNM 7th)II.Results of endoscopically treated patients in 2012TableDetails of endoscopic treatment for curative intentTableComplications of EMR/ESDTablePathological depth of tumor invasion of EMR/ESD specimensFigureSurvival of patients treated with EMR/ESDFigureSurvival of patients treated with EMR/ESD according to the pathological depth of tumor invasion, pT (UICC TNM 7th)FigureSurvival of patients treated with EMR/ESD according to the lymphatic and venous invasionIII.Results in patients treated with chemotherapy and/or radiotherapy in 2012TableDose of irradiation TableDose of irradiation FigureSurvival of patients treated with chemotherapy and/or radiotherapyFigureSurvival of patients treated with definitive chemoradiotherapy according to clinical stage (UICC TNM 7th)FigureSurvival of patients underwent radiotherapy alone according to clinical stage (UICC TNM 7th)IV.Results in patients who underwent esophagectomy in 2012TableTreatment modalities of esophagectomyTableTumor locationTableApproaches to tumor resectionTableVideo-assisted surgeryTableFields of lymph node dissection according to the location of the tumorTableReconstruction routeTableOrgans used for reconstructionTableHistological classificationTableDepth of tumor invasion, pT (JES 10th)TablePathological grading of lymph node metastasis, pN (JES 10th)TablePathological findings of lymph node metastasis, pN (UICC 7th)TablePathological findings of distant organ metastasis, pM (JES 10th)TableResidual tumorTableCauses of deathFigureSurvival of patients who underwent esophagectomyFigureSurvival of patients who underwent esophagectomy according to clinical stage (JES 10th)FigureSurvival of patients who underwent esophagectomy according to clinical stage (UICC 7th)FigureSurvival of patients who underwent esophagectomy according to the depth of tumor invasion, pT (JES 10th)FigureSurvival of patients who underwent esophagectomy according to lymph node metastasis, pN (JES 10th)FigureSurvival of patients who underwent esophagectomy according to lymph node metastasis, pN (UICC 7th)FigureSurvival of patients who underwent esophagectomy according to pathological stage (JES 10th)FigureSurvival of patients who underwent esophagectomy according to pathological stage (UICC TNM 7th)FigureSurvival of patients who underwent esophagectomy according to residual tumor (R)Institution-registered cases in 2012Tables Tables Tables Tables 22