Immunohistochemistry of thyroid hormones as a functional parameter in thyroid pathology

The authors study by means of immunoperoxidase method the pattern of thyroglobulin, triiodothyronine and thyroxine distribution in 58 cases of thyroid disorders: 15 euthyroid goiters, 10 Graves' disease, 7 Hashimoto's thyroiditis, 11 folliculo-papillary carcinomas (6 primary tumors and 5 lymph node metastases), 8 follicular carcinomas, 4 anaplastic carcinomas and 3 medullary carcinomas. Thyroglobulin, triiodothyronine and thyroxine were present in most of the thyroid disorders, excepting anaplastic and medullary carcinomas. Thyroglobulin and thyroxine were localized both in the follicular epithelium and in the colloid, whereas triiodothyronine was present especially in the follicular cells. The thyroid hormones distribution in benign lesions is rather similar. In carcinomas, the pattern of thyroglobulin, triiodothyronine and thyroxine is more heterogeneous, but generally the triiodothyronine distribution is similar to that of thyroglobulin. In some carcinomas, triiodothyronine and thyroxine showed a weak or negative immunostaining. The immunoperoxidase method is a valuable tool in the study of functional disturbances in the thyroid pathology and in the diagnosis of thyroid carcinoma metastases as well. Positive thyroid hormones staining clearly indicates the thyroid origin of metastases.     

Histopathologic observations on thyroid carcinoma with amyloid stroma.

The authors describe histopathologic aspects in ten cases of thyroid carcinoma with amyloid stroma. This form of cancer represents 3.4% of all the thyroid cancer cases in the last five years in the Institute of Endocrinology. Microscopic examination reveals a polymorphous picture alongside with amyloid deposits. The authors attribute to the tumor cells an important role in producing thyroid amyloidosis.     

Targeted Next Generation Sequencing as a Reliable Diagnostic Assay for the Detection of Somatic Mutations in Tumours Using Minimal DNA Amounts from Formalin Fixed Paraffin Embedded Material

Background

Targeted Next Generation Sequencing (NGS) offers a way to implement testing of multiple genetic aberrations in diagnostic pathology practice, which is necessary for personalized cancer treatment. However, no standards regarding input material have been defined. This study therefore aimed to determine the effect of the type of input material (e.g. formalin fixed paraffin embedded (FFPE) versus fresh frozen (FF) tissue) on NGS derived results. Moreover, this study aimed to explore a standardized analysis pipeline to support consistent clinical decision-making.

Method

We used the Ion Torrent PGM sequencing platform in combination with the Ion AmpliSeq Cancer Hotspot Panel v2 to sequence frequently mutated regions in 50 cancer related genes, and validated the NGS detected variants in 250 FFPE samples using standard diagnostic assays. Next, 386 tumour samples were sequenced to explore the effect of input material on variant detection variables. For variant calling, Ion Torrent analysis software was supplemented with additional variant annotation and filtering.

Results

Both FFPE and FF tissue could be sequenced reliably with a sensitivity of 99.1%. Validation showed a 98.5% concordance between NGS and conventional sequencing techniques, where NGS provided both the advantage of low input DNA concentration and the detection of low-frequency variants. The reliability of mutation analysis could be further improved with manual inspection of sequence data.

Conclusion

Targeted NGS can be reliably implemented in cancer diagnostics using both FFPE and FF tissue when using appropriate analysis settings, even with low input DNA.     

<Emphasis Type="Italic">Usnea antarctica</Emphasis>, an important Antarctic lichen,is vulnerable to aspects of regional environmental change

Studies of cryptogam responses to climate change in the polar regions are scarce because these slow-growing organisms require long-term monitoring studies. Here, we analyse the response of a lichen and moss community to 10 years of passive environmental manipulation using open-top chambers (OTCs) in the maritime Antarctic region. Cover of the dominant lichen Usnea antarctica declined by 71 % in the OTCs. However, less dominant lichen species showed no significant responses except for an increase in Ochrolechia frigida, which typically covered dying lichen and moss vegetation. There were no detectable responses in the moss or associated micro-arthropod communities to the influence of the OTCs. Based on calculated respiration rates, we hypothesise that the decline of U. antarctica was most likely caused by increased net winter respiration rates (11 %), driven by the higher temperatures and lower light levels experienced inside the OTCs as a result of greater snow accumulation. During summer, U. antarctica appears unable to compensate for this increased carbon loss, leading to a negative carbon balance on an annual basis, and the lichen therefore appears to be vulnerable to such climate change simulations. These findings indicate that U. antarctica dominated fell-fields may change dramatically if current environmental change trends continue in the maritime Antarctic, especially if associated with increases in winter snow depth or duration.     

The Terneuzen Birth Cohort: BMI Changes between 2 and 6 Years Correlate Strongest with Adult Overweight

Background

Complications of overweight amplify with age, and irreversible damage already exists in young persons. Identifying the most sensitive age interval(s) for adult overweight is relevant for primary prevention. The aim of the study was to assess the relative contribution of body mass index (BMI) changes between 0 and 18 years to adult overweight, and to identify the earliest critical growth period.

Methods and Findings

Data from 762 subjects in the Terneuzen Birth Cohort with an average of 21 growth measurements per subject from birth until 18 years were used. The main outcome measure was the BMI standard deviation score (SDS) at young adulthood. For each subject BMI SDS was fitted by a piecewise linear model at eight different ages and correlated to adult BMI SDS. The age intervals in between are considered critical according to three criteria, tested by respectively Students'' t-tests, multiple linear regression analyses and Pearson''s correlation tests. In the age intervals 4 months(m) -1 year(y), 2–6 y, 6–10 y and 10–18 y the BMI SDS change differs between adults with and without overweight (P≤0.001). The age intervals 2–6 y and 10–18 y also meet the second criterion, implying that the BMI change during this period has a predictive value for adult BMI SDS in addition to BMI SDS at the end of the period. The largest rise in correlation between estimated BMI SDS and measured adult BMI SDS occurs during the period 2–6 y (from 0.36 to 0.63), which results in a high sensitivity (0.6) and specificity (0.8) by the age of 6 y.

Conclusions/Significance

The age interval from 2 y to 6 y is the earliest and most critical growth period for adult overweight. Therefore, primary prevention of adult overweight seems most likely to be successful if targeted at this specific age interval. By identifying those with an upwards centile crossing between 2 and 6 years, the development towards adult overweight might be reversed.     

Historical and contemporary gene dispersal in wild carrot (Daucus carota ssp. carota) populations

Background and Aims

Wild carrot is the ancestor of cultivated carrot and is the most important gene pool for carrot breeding. Transgenic carrot may be released into the environment in the future. The aim of the present study was to determine how far a gene can disperse in wild carrot populations, facilitating risk assessment and management of transgene introgression from cultivated to wild carrots and helping to design sampling strategies for germplasm collections.

Methods

Wild carrots were sampled from Meijendel and Alkmaar in The Netherlands and genotyped with 12 microsatellite markers. Spatial autocorrelation analyses were used to detect spatial genetic structures (SGSs). Historical gene dispersal estimates were based on an isolation by distance model. Mating system and contemporary pollen dispersal were estimated using 437 offspring of 20 mothers with different spatial distances and a correlated paternity analysis in the Meijendel population.

Key Results

Significant SGSs are found in both populations and they are not significantly different from each other. Combined SGS analysis indicated significant positive genetic correlations up to 27 m. Historical gene dispersal σ_g and neighbourhood size N_b were estimated to be 4–12 m [95 % confidence interval (CI): 3–25] and 42–73 plants (95 % CI: 28–322) in Meijendel and 10–31 m (95 % CI: 7–∞) and 57–198 plants (95 % CI: 28–∞) in Alkmaar with longer gene dispersal in lower density populations. Contemporary pollen dispersal follows a fat-tailed exponential-power distribution, implying pollen of wild carrots could be dispersed by insects over long distance. The estimated outcrossing rate was 96 %.

Conclusions

SGSs in wild carrots may be the result of high outcrossing, restricted seed dispersal and long-distance pollen dispersal. High outcrossing and long-distance pollen dispersal suggest high frequency of transgene flow might occur from cultivated to wild carrots and that they could easily spread within and between populations.     

F1F0 ATP synthase subunit c is a substrate of the novel YidC pathway for membrane protein biogenesis

The Escherichia coli YidC protein belongs to the Oxa1 family of membrane proteins that have been suggested to facilitate the insertion and assembly of membrane proteins either in cooperation with the Sec translocase or as a separate entity. Recently, we have shown that depletion of YidC causes a specific defect in the functional assembly of F1F0 ATP synthase and cytochrome o oxidase. We now demonstrate that the insertion of in vitro-synthesized F1F0 ATP synthase subunit c (F0c) into inner membrane vesicles requires YidC. Insertion is independent of the proton motive force, and proteoliposomes containing only YidC catalyze the membrane insertion of F0c in its native transmembrane topology whereupon it assembles into large oligomers. Co-reconstituted SecYEG has no significant effect on the insertion efficiency. Remarkably, signal recognition particle and its membrane-bound receptor FtsY are not required for the membrane insertion of F0c. In conclusion, a novel membrane protein insertion pathway in E. coli is described in which YidC plays an exclusive role.     

The restriction of zoonotic PERV transmission by human APOBEC3G

The human APOBEC3G protein is an innate anti-viral factor that can dominantly inhibit the replication of some endogenous and exogenous retroviruses. The prospects of purposefully harnessing such an anti-viral defense are under investigation. Here, long-term co-culture experiments were used to show that porcine endogenous retrovirus (PERV) transmission from pig to human cells is reduced to nearly undetectable levels by expressing human APOBEC3G in virus-producing pig kidney cells. Inhibition occurred by a deamination-independent mechanism, likely after particle production but before the virus could immortalize by integration into human genomic DNA. PERV inhibition did not require the DNA cytosine deaminase activity of APOBEC3G and, correspondingly, APOBEC3G-attributable hypermutations were not detected. In contrast, over-expression of the sole endogenous APOBEC3 protein of pigs failed to interfere significantly with PERV transmission. Together, these data constitute the first proof-of-principle demonstration that APOBEC3 proteins can be used to fortify the innate anti-viral defenses of cells to prevent the zoonotic transmission of an endogenous retrovirus. These studies suggest that human APOBEC3G-transgenic pigs will provide safer, PERV-less xenotransplantation resources and that analogous cross-species APOBEC3-dependent restriction strategies may be useful for thwarting other endogenous as well as exogenous retrovirus infections.