pp 209-218 Article
Approximately 98% of patients with multiple endocrine neoplasia type 2A (MEN 2A) have an identifiable RETmutation. Prophylactic or early total thyroidectomy or pheochromocytoma/parathyroid removal in patients can bepreventative or curative and has become standard management. The general strategy for RET screening on familymembers at risk is to sequence the most commonly affected exons and, if negative, to extend sequencing to additionalexons. However, different families with MEN 2A due to the same RET mutation often have significant variability inthe clinical exhibition of disease and aggressiveness of the MTC, which implies additional genetic loci exsit beyondRET coding region. Whole genome sequencing (WGS) greatly expands the breadth of screening from genes associatedwith a particular disease to the whole genome and, potentially, all the information that the genome containsabout diseases or traits. This is presumably due to additive effect of disease modifying factors. In this study, weperformed WGS on a typical Chinese MEN 2A proband and identified the pathogenic RET p.C634R mutation. Wealso identified several neutral variants within RET and pheochromocytoma-related genes. Moreover, we found severalinteresting structural variants including genetic deletions (RSPO1, OVCH2 and AP3S1, etc.) and fusion transcripts(FSIP1-BAZ2A, etc.).
pp 219-230 Article
The present study aims to investigate the role of radiation sensitive 52 (RAD52) and high-affinity DNA binding factor1 (HDF1) DNA repair genes on the life span of budding yeasts during chronological aging. Wild type (wt) and rad52,hdf1, and rad52 hdf1 mutant Saccharomyces cerevisiae strains were used. Chronological aging and survival assayswere studied by clonogenic assay and drop test. DNA damage was analyzed by electrophoresis after phenol extraction.Mutant analysis, colony forming units and the index of respiratory competence were studied by growing on dextroseand glycerol plates as a carbon source. Rad52 and rad52 hdf1 mutants showed a gradual decrease in surviving fractionin relation to wt and hdf1 mutant during aging. Genomic DNA was spontaneously more degraded during aging,mainly in rad52 mutants. This strain showed an increased percentage of revertant colonies. Moreover, all mutantsshowed a decrease in the index of respiratory competence during aging. The inactivation of RAD52 leads to prematurechronological aging with an increase in DNA degradation and mutation frequency. In addition, RAD52 and HDF1contribute to maintain the metabolic state, in a different way, during chronological aging. The results obtained couldhave important implications in the chronobiology of aging.
pp 231-244 Article
Intracellular trafficking and localization studies of spike protein from SARS and OC43 showed that SARS spikeprotein is localized in the ER or ERGIC compartment and OC43 spike protein is predominantly localized in thelysosome. Differential localization can be explained by signal sequence. The sequence alignment using Clustal Wshows that the signal sequence present at the cytoplasmic tail plays an important role in spike protein localization. Aunique GYQEL motif is identified at the cytoplasmic terminal of OC43 spike protein which helps in localization in thelysosome, and a novel KLHYT motif is identified in the cytoplasmic tail of SARS spike protein which helps in ER orERGIC localization. This study sheds some light on the role of cytoplasmic tail of spike protein in cell-to-cell fusion,coronavirus host cell fusion and subsequent pathogenicity.
pp 245-250 Article
Capsinoids are non-pungent analogues of capsaicinoids in pepper (Capsicum spp). The absence of pungency, inaddition to their biological activities similar to that of capsaicinoids such as anti-inflammatory, antimicrobial, andantioxidant properties, makes capsinoids an excellent option for increasing use in human and animal nutrition, as wellas health and pharmaceutical industries. There are only few sources of pepper producing capsinoids, and one of them(accession 509–45-1), Capsicum annuum L., is a potential source for increasing capsinoids content using strategies ascontrolled elicitation during plant production in the greenhouse. In this research we evaluated the effect of weekly andone-day-before-harvest foliar applications of hydrogen peroxide, salicylic acid and a xyloglucan oligosaccharide onthe concentration of capsiate in fruits of this pepper accession, as well as the gene expression of phenylalanineammonia-lyase (pal), putative aminotransferase (pamt), capsaicin synthase (at3) and β-keto acyl synthase (kas).Results showed that the two tested concentrations of H2O2 significantly increased capsiate content and geneexpression associated with capsaicinoids (pamt, at3 and kas) and the phenylpropanoids (pal) pathways. Plant yieldwas not affected using this induction strategy. Our results indicated that the pre-harvest and weekly application ofhydrogen peroxide and xyloglucan oligosaccharide improved production of capsiate in C. annuum L.
pp 251-263 Article
Endometriosis is a common gynaecological disorder of unknown aetiology. Among the several factors, estrogen hasbeen implicated as a causative factor in endometriosis. In the present study using mouse model, we assessed the role ofestrogen in the initial implantation and growth of endometrium in ectopic locations. Uterine tissues from greenfluorescent protein (GFP) mice were transplanted in to the peritoneum of wild type mice in presence and absence ofestrogen. As compared to untreated controls, the implantation of uterine tissue at ectopic locations was higher whenestrogen was administered to both host and donor animals. However, this effect was not sustained as lesions regressedwithin 14 days of treatment. Irrespective of the treatment, peritoneal adipose was the most preferred site of lesionestablishment. The lesions did not have typical features of the endometriosis (presence of glands and stroma) even afterestrogen treatment and the ectopic tissue underwent regression by apoptosis irrespective of treatment. Since estrogenpromotes implantation of endometrial tissue to ectopic locations but failure of these ectopic lesions to grow and sustaineven in high estrogenic environment we propose that estrogen is necessary but not sufficient to sustain endometriosis.
pp 265-274 Article
The only way to perceive the real clinical course of disease and the prognostic significance of potential biomarkers is follow-up of patients who did not receive any kind of adjuvant therapy. Many studies have confirmed high levels ofinterleukin 8 (IL8) in HER2-enriched and basal-like (ER–) primary breast tumours, but less is known about thesignificance of IL8 in hormone-dependent breast cancer. The aim of this study was to evaluate the prognostic significance of IL8 and clinicopathological parameters in hormone-dependent breast cancer, and to examine possible associations between them that might imply possible biological dependence. The study included 91 early-stage breastcancer patients with detectable levels of hormone receptors (ER>0, PR>0). None of the patients received adjuvanttherapy according to valid protocol at that time. HER2 status was determined on paraffin-embedded tumour tissue sections by CISH. IL8 levels were determined by ELISA in cytosol tumour extracts of 65 patients with long-term follow-up (144 months). Nonparametric statistical tests were used for data analyses. Patients with low IL8 levels(M<88.8 pg/mg) had significantly longer relapse-free survival (RFS) compared to patients with high IL8 levels(M≥88.82 pg/mg) (Log rank test, p=0.002). Patients with ERhighIL8low phenotype had significantly longer RFScompared to those with ERhighIL8high and ERlowIL8high phenotypes (p=0.04 and p=0.02, respectively); patientswith PRlowIL8low phenotype had significantly longer RFS compared to those with PRlowIL8high and PRhighIL8-high phenotypes (p=0.003 and p=0.02, respectively); patients with HER2-IL8low phenotype had significantly longerRFS compared to those with HER2-IL8high and HER2+IL8high phenotypes (p=0.01 and p=0.02, respectively). Ourresults indicate significant contribution of IL8 on survival of hormone-dependent early-stage breast cancer patientsand association with established parameters such as ER/PR and HER2.
pp 275-283 Article
Arginase II is a mitochondrial enzyme that catalyses the hydrolysis of L-arginine into urea and ornithine. It is presentin other extra-hepatic tissues that lack urea cycle. Therefore, it is plausible that arginase II has a physiological roleother than urea cycle which includes polyamine, proline, glutamate synthesis and regulation of nitric oxide production.The high expression of arginase II in kidney, among extrahepatic tissues, might have an important role associatedwith kidney functions. The present study is aimed to determine the age-associated alteration in the activity andexpression of arginase II in the kidney of mice of different ages. The effect of dietary restriction to modulate the agedependentchanges of arginase II was also studied. Results showed that renal arginase II activity declines significantlywith the progression of age (p<0.01 and p<0.001 in 6- and 18-month-old mice, respectively as compared to 2-monthold mice) and is due to the reduction in its protein as well as the mRNA level (p<0.001 in both 6- and 18-month-oldmice as compared to 2-month-old mice). Long-term dietary restriction for three months has significantly up-regulatedarginase II activity and expression level in both 2- and 18-month-old mice (p<0.01 and p<0.001, respectively ascompared to AL group). These findings clearly indicate that the reducing level of arginase II during aging might havean impact on the declining renal functions. This age-dependent down-regulation of arginase II in the kidney can beattenuated by dietary restriction which may help in the maintenance of such functions.
pp 285-297 Article
Spermatogenesis, involving multiple transit amplification divisions and meiosis, occurs within an enclosure formed bytwo somatic cells. As the cohort of germline cells divide and grow, the surface areas of the somatic cells expandmaintaining a tight encapsulation throughout the developmental period. Correlation between the somatic cell growthand germline development is unclear. Here, we report standardization of a quantitative assay developed for estimatingthe somatic roles of target molecules on germline division and differentiation in Drosophila testis. Using the assay, westudied the somatic roles of phosphatidylinositol-3-kinase (PI3K). It revealed that the expression of PI3KDN is likelyto facilitate the early germline development at all stages, and an increase in the somatic PI3K activity during the earlystages delays the transition to spermatocyte stage. Together, these results suggest that somatic cell growth plays animportant role in regulating the rate of germline development.
pp 299-309 Article
We investigated the efficacy and safety of ultrasound (US)-targeted microbubble (MB) destruction (UTMD)-enhanceddelivery of monomethoxypoly(ethylene glycol)-poly(lactic-co-glycolic acid)-poly-L-lysine (mPEG-PLGA-PLL)nanoparticles (NPs) loading Cy3-labelled platelet-derived growth factor BB (PDGF-BB) siRNA to rat retinain vivo. Eighty Wistar rats were divided into five groups (G). The right eyes, respectively, received an intravitrealinjection as follows: normal saline (NS) (G1), NPs and NS (G2), NPs and MBs (G3), NPs and NS (G4) and NPs andMBs (G5). In G4 and G5, the eyes were exposed to US for 5 mins. Twenty-four hours after transfection, the uptakeand distribution of Cy3-labelled siRNA in rat retina were observed by fluorescent microscope. The percentage of Cy3-labelled siRNA-positive cells was evaluated by flow cytometer. The levels of PDGF‐BB mRNA in retinal pigmentepithelium (RPE) cells and secreted PDGF‐BB proteins were also measured. Hematoxylin and eosin staining andfrozen sections were used to observe tissue damage. Our results showed that the number of Cy3-labelled siRNApositivecells in G5 was significantly higher than those of the other groups (P<0.05 for all comparisons). Themaximum efficiency of siRNA uptake in neural retina was 18.22±1.67%. In G4 and G5, a small number of Cy3-labelled siRNA-positive cells were also detected in the pigmented cell layer of the retina. NPs loading siRNAdelivered with UTMD could more effectively down-regulate the mRNA and protein expression of PDGF‐BB thanNPs plus US (P=0.014 and P=0.007, respectively). Histology showed no evident tissue damage after UTMDmediatedNPs loading siRNA transfection. UTMD could be used safely to enhance the delivery of mPEG-PLGAPLLNPs loading siRNA into rat retina.
pp 311-319 Article
We investigated whether maternal over-nutrition during pregnancy and lactation affects the offspring’s lipidmetabolism at weaning by assessing liver lipid metabolic gene expressions and analysing its mechanisms on thedevelopment of metabolic abnormalities. Female Sprague–Dawley rats were fed with standard chow diet (CON)or high-fat diet (HFD) for 8 weeks, and then continued feeding during gestation and lactation. The offspringwhose dams were fed with HFD had a lower birth weight but an increased body weight with impaired glucosetolerance, higher serum cholesterol, and hepatic steatosis at weaning. Microarray analyses showed that there were120 genes differently expressed between the two groups. We further verified the results by qRT-PCR. Significantincrease of the lipogenesis (Me1, Scd1) gene expression was found in HFD (P<0.05), and up-regulated expressionof genes (PPAR-α, Cpt1α, Ehhadh) involved in β-oxidation was also observed (P<0.05), but the Acsl3 gene wasdown-regulated (P<0.05). Maternal over-nutrition could not only primarily induce lipogenesis, but also promotelipolysis through an oxidation pathway as compensation, eventually leading to an increased body weight,impaired glucose tolerance, elevated serum cholesterol and hepatic steatosis at weaning. This finding may providesome evidence for a healthy maternal diet in order to reduce the risk of metabolic diseases in the early life of theoffspring.
pp 321-331 Review
While living collections are critical for biological research, support for these foundational infrastructure elements isinconsistent, which makes quality control, regulatory compliance, and reproducibility difficult. In recent years, theEcological Society of America has hosted several National Science Foundation–sponsored workshops to explore andenhance the sustainability of biological research infrastructure. At the same time, the United States Culture CollectionNetwork has brought together managers of living collections to foster collaboration and information exchange withina specific living collections community. To assess the sustainability of collections, a survey was distributed tocollection scientists whose responses provide a benchmark for evaluating the resiliency of these collections. Amongthe key observations were that plant collections have larger staffing requirements and that living microbe collectionswere the most vulnerable to retirements or other disruptions. Many higher plant and vertebrate collections haveinstitutional support and several have endowments. Other collections depend on competitive grant support in an era ofintense competition for these resources. Opportunities for synergy among living collections depend upon complementingthe natural strong engagement with the research communities that depend on these collections with enhancedinformation sharing, communication, and collective action to keep them sustainable for the future. External efforts byfunding agencies and publishers could reinforce the advantages of having professional management of researchresources across every discipline.
pp 333-344 Review
Dynamin superfamily proteins comprising classical dynamins and related proteins are membrane remodelling agentsinvolved in several biological processes such as endocytosis, maintenance of organelle morphology and viralresistance. These large GTPases couple GTP hydrolysis with membrane alterations such as fission, fusion ortubulation by undergoing repeated cycles of self-assembly/disassembly. The functions of these proteins are regulatedby various post-translational modifications that affect their GTPase activity, multimerization or membrane association.Recently, several reports have demonstrated variety of such modifications providing a better understanding of themechanisms by which dynamin proteins influence cellular responses to physiological and environmental cues. In thisreview, we discuss major post-translational modifications along with their roles in the mechanism of dynaminfunctions and implications in various cellular processes.
pp 345-353 Review
FSHD region gene 1 (FRG1), as the name suggests, is the primary candidate gene for fascioscapulohumeral musculardystrophy disease. It seemingly affects muscle physiology in normal individuals but in FSHD, where it is found to behighly upregulated, might be involved in disruption of face, scapula and humeral skeletal muscle. Literature on FRG1,reviewed from 1996 to 2016, reveals that it is primarily associated with muscle development and maintenance.Approximately 75% of FSHD patients also show vascular abnormalities indicating that FRG1 might have some part toplay in these abnormalities. Research involving vasculature in X. laevis larvae shows that FRG1 positively affectsnormal vasculature. Few of the well-established angiogenic regulators seem to get affected by abnormal expressionlevel of FRG1. Its primary localization in sub nuclear structures like Cajal bodies and nuclear speckles indicatesregulation of the above-mentioned factors by transcriptional and post-transcriptional machineries, but in-depth studiesneed to be done to conclude a clear statement. In this review, we have attempted to present all the work done onFRG1, all the lacunas which need to be unraveled, and hypothesized a model for our readers to get an insight into itsmolecular mechanism.