Needle track seeding of papillary thyroid carcinoma from fine needle aspiration biopsy. A case report

BACKGROUND: Dissemination of tumor cells from needle biopsy has been observed in a wide range of tumor types. Fine needle aspiration (FNA) biopsy has become accepted as the first-line test in the evaluation of thyroid nodules. Local recurrence of thyroid cancer from needle track seeding is an extremely rare complication of thyroid FNA. CASE: A 59-year-old woman developed local recurrence of papillary thyroid carcinoma three years after FNA of the primary cancer. Local metastases developed in the skin and sternocleidomastoid muscle. The location of the recurrent cancer and the linear relationship of the metastases indicated that local recurrence was due to needle track seeding at the time of FNA. CONCLUSION: Needle track seeding has been recognized as a possible, albeit rare, complication of FNA of thyroid cancer. Although proper FNA technique can reduce the potential for needle track seeding, its occurrence is an unavoidable complication of FNA evaluation of thyroid malignancies.     

Utility of thiol-cross-linked fluorescent dye labeled terminators for DNA sequencing

Dipivaloyl-5-carboxyfluorescein N-hydroxysuccinimidyl ester 1 and 5-propargylamino-2',3'-dideoxyuridine triphosphate 5 were modified with maleimide, haloacetamide, and sulfhydryl reactive functional groups to participate in cross-conjugation reactions via sulfide bonds to generate fluorescently labeled, thioether cross-conjugated terminators 10 and 11. Their DNA sequencing potential was compared with an amide cross-conjugated terminator 13, synthesized by directly coupling 5-carboxyfluorescein NHS ester with 18-ddUTP 9. These terminators (10, 11, and 13) in combination with the Thermo Sequenase II DNA polymerase, in thermal cycle sequencing experiments, revealed that the thioether cross-conjugated terminator 10 and amide cross-conjugated terminator 13 served as good terminating substrates, generating satisfactory single-color gel images and electropherograms, while the other thioether cross-conjugated and maleimide derived one 11 underwent unexpected pH and temperature induced decomposition without showing fluorescent signatures for incorporation.     

Genomic organization and comparative sequence analysis of the mouse and human FRS2, FRS3 genes

The signaling adapter proteins FRS2 and FRS3 are implicated in the transmission of extracellular signals from nerve growth factor (NGF) or fibroblast growth factor (FGF) receptors to the Ras/mitogen-activated protein kinase signaling cascade. This study presents the genomic sequence and exon-intron organization of the mouse FRS2 and FRS3 loci as well as their evolutionary conservation with their human counterparts. Both FRS2 and FRS3 contain 5 coding exons spanning over 7 kb of genomic sequence with similar exon sizes and organization. Comparative genomic sequence analyses show a highly conserved genomic organization between mouse and human in both FRS2 and FRS3 genes. Non-coding sequences, highly conserved between mouse and human, were identified in the FRS3 introns that may potentially function as regulatory elements. To assay potential differences in their patterns of expression, RT-PCR analysis was used to assay FRS2 and FRS3 expression in the developing embryo and neural tube (NT) during the time of neurogenesis.     

Late gestational changes in sympathomimetic sensitivity in primagravid rabbit ligaments

The adrenoceptor profile of blood vessels supplying the medial collateral ligament (MCL) of virgin and primagravid (day 29 of pregnancy) rabbit knees was examined. Topical bolus administration of the alpha1-adrenoceptor agonists methoxamine and phenylephrine, and the alpha2-adrenoceptor agonist clonidine, to exposed knee joints resulted in a dose-dependent vasoconstriction of ligamentous vessels. The rank order of potency for the alpha-agonists was found to be phenylephrine > methoxamine > clonidine. Dobutamine, which acts on beta1-adrenoceptors, and ritodrine, which is a beta2-agonist, imparted a mild vasodilatatory effect on MCL blood vessels with the efficacy of ritodrine being greater than that of dobutamine. In primagravid rabbits, the constrictor effects of methoxamine and phenylephrine were significantly attenuated compared with virgin control animals (P < 0.0001), whereas clonidine-mediated vasoconstriction was unaltered in the gravid animal (P = 0.3957). With respect to beta-adrenoceptor activity, the dilatational effect of dobutamine was the same as in controls (P = 0.5294), while ritodrine vasoactivity was completely abolished in primagravid knees (P < 0.005). These findings suggest that the adrenergic control of rabbit MCL blood flow is predominantly mediated by postjunctional alpha1 and beta2-adrenoceptors. Pregnancy leads to either downregulation and (or) desensitisation of alpha1 and beta2-adrenoceptors in the ligament which could disrupt normal joint homeostasis.     

Extremely Low Microsatellite Diversity but Distinct Population Structure in a Long-Lived Threatened Species,the Australian Lungfish Neoceratodus forsteri (Dipnoi)

The Australian lungfish is a unique living representative of an ancient dipnoan lineage, listed as ‘vulnerable’ to extinction under Australia’s Environment Protection and Biodiversity Conservation Act 1999. Historical accounts indicate this species occurred naturally in two adjacent river systems in Australia, the Burnett and Mary. Current day populations in other rivers are thought to have arisen by translocation from these source populations. Early genetic work detected very little variation and so had limited power to answer questions relevant for management including how genetic variation is partitioned within and among sub-populations. In this study, we use newly developed microsatellite markers to examine samples from the Burnett and Mary Rivers, as well as from two populations thought to be of translocated origin, Brisbane and North Pine. We test whether there is significant genetic structure among and within river drainages; assign putatively translocated populations to potential source populations; and estimate effective population sizes. Eleven polymorphic microsatellite loci genotyped in 218 individuals gave an average within-population heterozygosity of 0.39 which is low relative to other threatened taxa and for freshwater fishes in general. Based on F _ST values (average over loci = 0.11) and STRUCTURE analyses, we identify three distinct populations in the natural range, one in the Burnett and two distinct populations in the Mary. These analyses also support the hypothesis that the Mary River is the likely source of translocated populations in the Brisbane and North Pine rivers, which agrees with historical published records of a translocation event giving rise to these populations. We were unable to obtain bounded estimates of effective population size, as we have too few genotype combinations, although point estimates were low, ranging from 29 - 129. We recommend that, in order to preserve any local adaptation in the three distinct populations that they be managed separately.     

Induced root-secreted phenolic compounds as a belowground plant defense

Rhizosphere is the complex place of numerous interactions between plant roots, microbes and soil fauna. Whereas plant interactions with aboveground organisms are largely described, unravelling plant belowground interactions remains challenging. Plant root chemical communication can lead to positive interactions with nodulating bacteria, mycorriza or biocontrol agents or to negative interactions with pathogens or root herbivores. A recent study¹ suggested that root exudates contribute to plant pathogen resistance via secretion of antimicrobial compounds. These findings point to the importance of plant root exudates as belowground signalling molecules, particularly in defense responses. In our report,² we showed that under Fusarium attack the barley root system launched secretion of phenolic compounds with antimicrobial activity. The secretion of de novo biosynthesized t-cinnamic acid induced within 2 days illustrates the dynamic of plant defense mechanisms at the root level. We discuss the costs and benefits of induced defense responses in the rhizosphere. We suggest that plant defense through root exudation may be cultivar dependent and higher in wild or less domesticated varieties.Key words: root exudates, plant defense, t-cinnamic acid, fusarium, induced defensePlants grow and live in very complex and changing ecosystems. Because plants lack the mobility to escape from attack by pathogens or herbivores, they have developed constitutive and in addition inducible defenses that are triggered by spatiotemporally dynamic signaling mechanisms. These defenses counteract the aggressor directly via toxins or defense plant structures or indirectly by recruitment of antagonists of aggressors. Whereas induced defenses are well described in aboveground interactions, evidence of the occurrence of such mechanisms in belowground interactions remains limited. The biosynthesis of a defensive molecule could be both constitutive and inducible with a low level of a preformed pool (Fig. 1). In addition, upon encounter of an attacking organism, those levels could be induced to rise locally to a high level of active compound that is able to disarm the pathogen.^2,3 Only a few examples show that root exudates play a role in induced plant defense. Hairy roots of Ocimum basilicum secrete rosmarinic acid only when challenged by the pathogenic fungus Pythium ultimum.⁴ Wurst et al.⁵ reported on the induction of irridoid glycosides in root exudates of Plantago lanceolata in presence of nematodes. In vivo labelling experiments² with ¹³CO₂ showed the induction of phenolic compounds secreted by barley roots after Fusarium graminearum infection and the de novo biosynthesis of root secreted t-cinnamic acid within 2 days. These results show that the pool of induced t-cinnamic acid originated from both pre-formed and newly formed carbon pools (Fig. 1), highlighting a case of belowground induced defense inside and outside the root system.Open in a separate windowFigure 1Suggested mechanisms for the induction of root defense exudates in barley in response to Fusarium attack. Upon pathogen attack by Fusarium, the initial preformed pool of phenolic compounds is increased by the addition of inducible, de novo biosynthesized t-cinnamic acid. Both, the preformed pool and the de novo biosynthesized pool fuel the exudation of defense compounds from infected roots.The concept of fitness costs is frequently presented to explain the coexistence of both constitutive and induced defense.⁶ In the case of induced defense, resources are invested in defenses only when the plant is under attack. In the absence of an infection, plants can optimize allocation of their resources to reproduction and growth to compete with neighbours.⁷ Constitutive defenses are thought to be more beneficial when the probability of attack is high, whereas adjustable, induced defenses are more valuable to fight against an unpredictable pathogen. Non disturbed soil is a heterogeneous matrix where biodiversity is very high and patchy^8,9 and organism motility is rather restricted.¹⁰ As a consequence of the patchiness, belowground environment is expected to be favourable to selection for induced responses.¹¹ The absence of defense root exudates between two infections may form an unpredictable environment for soil pathogens and reduce the chance for adaptation of root attackers. Plants may also use escape strategies to reduce the effect of belowground pathogens. Henkes et al. (unpublished) showed that Fusarium-infected barley plants reduced carbon allocation towards infected roots within a day and increased allocation carbon to uninfected roots. These results illustrate how reallocation of carbon toward non infected root parts represents a way to limit the negative impact of root infection.We have demonstrated the potential of barley plants to defend themselves against soil pathogen by root exudation.² Even the barley cultivar ‘Barke’ used in our study, a modern cultivated variety, was able to launch defense machinery via exudation of antimicrobial compounds when infected by F. graminearum. We suggest that plant defense through root exudation might be cultivar dependent and perhaps higher in wild or less domesticated varieties. Taddei et al.¹² reported that constitutivelyproduced root exudates from a resistant Gladiolus cultivar inhibit spore germination of Fusarium oxysporum whereas root exudates from a susceptible cultivar do not affect F. oxysporum germination. Root exudates from the resistant cultivar contained higher amounts of aromaticphenolic compounds compared to the susceptible cultivar and these compounds may be responsible for the inhibition of spore germination. Metabolic profiling of wheat cultivars, ‘Roblin’ and ‘Sumai3’, respectively, susceptible and resistant to Fusarium Head Blight, showed that t-cinnamic acid was a discriminating factor responsible for resistance/defense function.¹³ Therefore it is likely that wild barley varieties hold higher defense capacities compare to cultivated varieties selected for high yield. In the future, plant breeders in organic and low-input farming could use root-system defense ability as new trait in varietal variation.     

Association between endometritis diagnosis using a novel intravaginal device and reproductive performance in dairy cattle

Endometritis reduces reproductive performance in dairy cattle. Diagnosis of endometritis is undertaken using a variety of techniques including vaginoscopy, manual examination, cytology and ultrasonography. The current studies compared a novel test device ("metricheck") that is inserted into the vagina with vaginoscopy and then examined the relationship between the metricheck test score at 35 days before the start of the seasonal breeding programme and subsequent reproductive performance. Cows (n = 191; Study 1) with a history of a peripartum disease were examined by both vaginoscopy and the metricheck device and any material viewed within the vagina (using vaginoscopy) or retrieved (by the metricheck device) was scored on a 0 (no material) to 5 (grossly purulent and with an odour) scale. Within each herd the order of examination was randomized with sequentially presented pairs of cows. All cows (n = 2793; Study 2) from nine herds were examined and scored using the metricheck device 35 days before the start of the seasonal breeding programme. All cows were pregnancy tested to determine date of conception. In Study 1, more cows were defined as infected (i.e. score > 1) following metricheck than vaginoscopic examination (60% versus 43%, respectively; P < 0.05) and the level of agreement between the two tests was moderate (kappa = 0.45). The metricheck device had a higher sensitivity, but lower specificity, than vaginoscopy. Endometritis (i.e. score > 1) was detected in 21.2% of cows examined in Study 2. The prevalence of endometritis varied among herds, declined with time postpartum (P < 0.05) and was higher in cows recorded as having a peripartum disease (P < 0.01). Cows diagnosed with endometritis were at higher risk of not being detected in oestrus before the start of breeding (P < 0.01), took longer to be inseminated after the start of the seasonal breeding programme (P < 0.01), had a lower first service conception rate (P < 0.01), had lower 56-day and final pregnancy rates (P < 0.05) and took longer to conceive than cows without endometritis (P < 0.05). It is concluded that examination with the metricheck device is more sensitive in detecting endometritis than vaginoscopy. Diagnosis of endometritis with the metricheck device was associated with poorer subsequent reproductive performance.     

Design, synthesis, and antiviral evaluation of some 3'-carboxymethyl-3'-deoxyadenosine derivatives

3'-Carboxymethyl-3'-deoxyadenosine derivatives were prepared from 2'-O-TBDMS-3'-[(ethoxycarbonyl)methyl]-3'-deoxyadenosine (1) via simple and efficient procedures. Conversion of 1 to its 5'-azido-5'-deoxy derivative 5 was accomplished via a novel one-pot method employing 5'-activation (TosCl) followed by efficient nucleophilic displacement with tetramethylguanidinium azide. Compound 5 was converted to 5'-[(N-methylcarbamoyl)amino] derivative 8 via one-pot reduction/acylation employing H(2)/Pd-C followed by treatment with p-nitrophenyl N-methylcarbamate. N(6)-phenylcarbamoyl groups were introduced by treatment with phenylisocyanate, and an efficient new method for lactonization of 2'-O-TBDMS-3'-[(ethoxycarbonyl)methyl]-3'-deoxyadenosines to give corresponding 2',3'-lactones was also developed. Target compounds were evaluated for anti-HIV and anti-HIV integrase activities, but were not active at the concentrations tested.     

Optimisation of a quantitative polymerase chain reaction-based strategy for the detection and quantification of human herpesvirus 6 DNA in patients undergoing allogeneic haematopoietic stem cell transplantation

Human herpesvirus 6 (HHV-6) may cause severe complications after haematopoietic stem cell transplantation (HSCT). Monitoring this virus and providing precise, rapid and early diagnosis of related clinical diseases, constitute essential measures to improve outcomes. A prospective survey on the incidence and clinical features of HHV-6 infections after HSCT has not yet been conducted in Brazilian patients and the impact of this infection on HSCT outcome remains unclear. A rapid test based on real-time quantitative polymerase chain reaction (qPCR) has been optimised to screen and quantify clinical samples for HHV-6. The detection step was based on reaction with TaqMan^® hydrolysis probes. A set of previously described primers and probes have been tested to evaluate efficiency, sensitivity and reproducibility. The target efficiency range was 91.4% with linearity ranging from 10-10⁶ copies/reaction and a limit of detection of five copies/reaction or 250 copies/mL of plasma. The qPCR assay developed in the present study was simple, rapid and sensitive, allowing the detection of a wide range of HHV-6 loads. In conclusion, this test may be useful as a practical tool to help elucidate the clinical relevance of HHV-6 infection and reactivation in different scenarios and to determine the need for surveillance.     

Association of 25-Hydroxyvitamin D status and genetic variation in the vitamin D metabolic pathway with FEV1 in the Framingham Heart Study

Background

Vitamin D is associated with lung function in cross-sectional studies, and vitamin D inadequacy is hypothesized to play a role in the pathogenesis of chronic obstructive pulmonary disease. Further data are needed to clarify the relation between vitamin D status, genetic variation in vitamin D metabolic genes, and cross-sectional and longitudinal changes in lung function in healthy adults.

Methods

We estimated the association between serum 25-hydroxyvitamin D [25(OH)D] and cross-sectional forced expiratory volume in the first second (FEV₁) in Framingham Heart Study (FHS) Offspring and Third Generation participants and the association between serum 25(OH)D and longitudinal change in FEV₁ in Third Generation participants using linear mixed-effects models. Using a gene-based approach, we investigated the association between 241 SNPs in 6 select vitamin D metabolic genes in relation to longitudinal change in FEV₁ in Offspring participants and pursued replication of these findings in a meta-analyzed set of 4 independent cohorts.

Results

We found a positive cross-sectional association between 25(OH)D and FEV₁ in FHS Offspring and Third Generation participants (P = 0.004). There was little or no association between 25(OH)D and longitudinal change in FEV₁ in Third Generation participants (P = 0.97). In Offspring participants, the CYP2R1 gene, hypothesized to influence usual serum 25(OH)D status, was associated with longitudinal change in FEV₁ (gene-based P < 0.05). The most significantly associated SNP from CYP2R1 had a consistent direction of association with FEV₁ in the meta-analyzed set of replication cohorts, but the association did not reach statistical significance thresholds (P = 0.09).

Conclusions

Serum 25(OH)D status was associated with cross-sectional FEV₁, but not longitudinal change in FEV₁. The inconsistent associations may be driven by differences in the groups studied. CYP2R1 demonstrated a gene-based association with longitudinal change in FEV₁ and is a promising candidate gene for further studies.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-015-0238-y) contains supplementary material, which is available to authorized users.