Genome-Wide and Locus Specific Alterations in CDC73/HRPT2-Mutated Parathyroid Tumors

Mutations in the hyperparathyroidism type 2 (HRPT2/CDC73) gene and alterations in the parafibromin protein have been established in the majority of parathyroid carcinomas and in subsets of parathyroid adenomas. While it is known that CDC73-mutated parathyroid tumors display specific gene expression changes compared to CDC73 wild-type cases, the molecular cytogenetic profile in CDC73-mutated cases compared to unselected adenomas (with an expected very low frequency of CDC73 mutations) remains unknown. For this purpose, nine parathyroid tumors with established CDC73 gene inactivating mutations (three carcinomas, one atypical adenoma and five adenomas) were analyzed for copy number alterations and loss of heterozygosity using array-comparative genomic hybridization (a-CGH) and single nucleotide polymorphism (SNP) microarrays, respectively. Furthermore, CDC73 gene promoter methylation levels were assessed using bisulfite Pyrosequencing. The panel included seven tumors with single mutation and three with double mutations of the CDC73 gene. The carcinomas displayed copy number alterations in agreement with previous studies, whereas the CDC73-mutated adenomas did not display the same pattern of alterations at loci frequently deleted in unselected parathyroid tumors. Furthermore, gross losses of chromosomal material at 1p and 13 were significantly (p = 0.012) associated with parathyroid carcinomas as opposed to adenomas. Quantitative PCR-based copy number loss regarding CDC73 was observed in three adenomas, while all the carcinomas were diploid or showed copy number gain for CDC73 gene. Hypermethylation of the CDC73 gene promoter was not observed. Our data could suggest that CDC73-mutated parathyroid adenomas exhibit a partly unique cytogenetic profile in addition to that of carcinomas and unselected adenomas. Furthermore, CDC73-mutated carcinomas displayed losses at 1p and 13 which are not seen in CDC73-mutated adenomas, making these regions of interest for further studies regarding malignant properties in tumors from CDC73-mutated cases. However, due to the small sample size, validation of the results in a larger cohort is warranted.     

Production of phosphatidylinositol 5‐phosphate via PIKfyve and MTMR3 regulates cell migration

Although phosphatidylinositol 5‐phosphate (PtdIns5P) is present in many cell types and its biogenesis is increased by diverse stimuli, its precise cellular function remains elusive. Here we show that PtdIns5P levels increase when cells are stimulated to move and we find PtdIns5P to promote cell migration in tissue culture and in a Drosophila in vivo model. First, class III phosphatidylinositol 3‐kinase, which produces PtdIns3P, was shown to be involved in migration of fibroblasts. In a cell migration screen for proteins containing PtdIns3P‐binding motifs, we identified the phosphoinositide 5‐kinase PIKfyve and the phosphoinositide 3‐phosphatase MTMR3, which together constitute a phosphoinositide loop that produces PtdIns5P via PtdIns(3,5)P₂. The ability of PtdIns5P to stimulate cell migration was demonstrated directly with exogenous PtdIns5P and a PtdIns5P‐producing bacterial enzyme. Thus, the identified phosphoinositide loop defines a new role for PtdIns5P in cell migration.     

The impact of simulated chronic nitrogen deposition on the biomass and N2-fixation activity of two boreal feather moss–cyanobacteria associations

Bryophytes achieve substantial biomass and play several key functional roles in boreal forests that can influence how carbon (C) and nitrogen (N) cycling respond to atmospheric deposition of reactive nitrogen (N_r). They associate with cyanobacteria that fix atmospheric N₂, and downregulation of this process may offset anthropogenic N_r inputs to boreal systems. Bryophytes also promote soil C accumulation by thermally insulating soils, and changes in their biomass influence soil C dynamics. Using a unique large-scale (0.1 ha forested plots), long-term experiment (16 years) in northern Sweden where we simulated anthropogenic N_r deposition, we measured the biomass and N₂-fixation response of two bryophyte species, the feather mosses Hylocomium splendens and Pleurozium schreberi. Our data show that the biomass declined for both species; however, N₂-fixation rates per unit mass and per unit area declined only for H. splendens. The low and high treatments resulted in a 29% and 54% reduction in total feather moss biomass, and a 58% and 97% reduction in total N₂-fixation rate per unit area, respectively. These results help to quantify the sensitivity of feather moss biomass and N₂ fixation to chronic N_r deposition, which is relevant for modelling ecosystem C and N balances in boreal ecosystems.     

Effects of High Doses of Cholecalciferol in Normal Subjects: A Randomized Double-Blinded,Placebo-Controlled Trial

Background

Vitamin D repletion with high doses of vitamin D is often recommended to patients and healthy subjects. The safety, especially concerning changes in urinary calcium excretion is of great importance.

Methods

In a double-blinded, placebo-controlled study in 40 healthy volunteers, we examined the changes in mineral metabolism during supplementation with 3000 IU of oral cholecalciferol daily during 4 months.

Results

Both 25(OH)vitamin D and 1,25(OH)₂vitamin D increased significantly in the active treated group as compared to the placebo group (186% versus 14% (P<0.001) and 28% versus – 8% (P<0.001)). No change was observed in urinary calcium excretion in the active group compared to the placebo group (P = 0.891). Fibroblast growth factor 23 increased significantly by 10% (P<0.018) in the active group. However, there was no difference in changes in FGF23 between treatment groups (P = 0.457).

Conclusion

High dose cholecalciferol significantly increases 25(OH)vitamin D and 1,25(OH)₂vitamin D levels compared to placebo. No changes in urinary calcium excretion or other measured components of the mineral metabolism were found between groups.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00952562","term_id":"NCT00952562"}}NCT00952562.     

Attempted Suicide in Bipolar Disorder: Risk Factors in a Cohort of 6086 Patients

Objective

Bipolar disorder is associated with high risk of self-harm and suicide. We wanted to investigate risk factors for attempted suicide in bipolar patients.

Method

This was a cohort study of 6086 bipolar patients (60% women) registered in the Swedish National Quality Register for Bipolar Disorder 2004–2011 and followed-up annually 2005–2012. Logistic regression was used to calculate adjusted odds ratios for fatal or non-fatal attempted suicide during follow-up.

Results

Recent affective episodes predicted attempted suicide during follow-up (men: odds ratio = 3.63, 95% CI = 1.76–7.51; women: odds ratio = 2.81, 95% CI = 1.78–4.44), as did previous suicide attempts (men: odds ratio = 3.93, 95% CI = 2.48–6.24; women: odds ratio = 4.24, 95% CI = 3.06–5.88) and recent psychiatric inpatient care (men: odds ratio = 3.57, 95% CI = 1.59–8,01; women: odds ratio = 2.68, 95% CI = 1.60–4.50). Further, those with many lifetime depressive episodes were more likely to attempt suicide. Comorbid substance use disorder was a predictor in men; many lifetime mixed episodes, early onset of mental disorder, personality disorder, and social problems related to the primary group were predictors in women.

Conclusion

The principal clinical implication of the present study is to pay attention to the risk of suicidal behaviour in bipolar patients with depressive features and more severe or unstable forms of the disorder.     

Turn structures in CGRP C-terminal analogues promote stable arrangements of key residue side chains.

The 37-amino acid calcitonin gene-related peptide (CGRP) is a potent endogenous vasodilator thought to be implicated in the genesis of migraine attack. CGRP antagonists may thus have therapeutic value for the treatment of migraine. The CGRP C-terminally derived peptide [D(31),P(34),F(35)]CGRP(27-37)-NH(2) was recently identified as a high-affinity hCGRP(1) receptor selective antagonist. Reasonable CGRP(1) affinity has also been demonstrated for several related analogues, including [D(31),A(34),F(35)]CGRP(27-37)-NH(2). In the study presented here, conformational and structural features in CGRP(27-37)-NH(2) analogues that are important for hCGRP(1) receptor binding were explored. Structure-activity studies carried out on [D(31),P(34),F(35)]CGRP(27-37)-NH(2) resulted in [D(31),P(34),F(35)]CGRP(30-37)-NH(2), the shortest reported CGRP C-terminal peptide analogue exhibiting reasonable hCGRP(1) receptor affinity (K(i) = 29.6 nM). Further removal of T(30) from the peptide's N-terminus greatly reduced receptor affinity from the nanomolar to micromolar range. Additional residues deemed critical for hCGRP(1) receptor binding were identified from an alanine scan of [A(34),F(35)]CGRP(28-37)-NH(2) and included V(32) and F(37). Replacement of the C-terminal amide in this same peptide with a carboxyl, furthermore, resulted in a greater than 50-fold reduction in hCGRP(1) affinity, thus suggesting a direct role for the amide moiety in receptor binding. The conformational properties of two classes of CGRP(27-37)-NH(2) peptides, [D(31),X(34),F(35)]CGRP(27-37)-NH(2) (X is A or P), were examined by NMR spectroscopy and molecular modeling. A beta-turn centered on P(29) was a notable feature consistently observed among active peptides in both series. This turn led to exposure of the critical T(30) residue to the surrounding environment. Peptides in the A(34) series were additionally characterized by a stable C-terminal helical turn that resulted in the three important residues (T(30), V(32), and F(37)) adopting consistent interspatial positions with respect to one another. Peptides in the P(34) series were comparatively more flexible at the C-terminus, although a large proportion of the [D(31),P(34),F(35)]CGRP(27-37)-NH(2) calculated conformers contained a gamma-turn centered on P(34). These results collectively suggest that turn structures at both the C-terminus and N-terminus of CGRP(27-37)-NH(2) analogues may help to appropriately orient critical residues (T(30), V(32), and F(37)) for hCGRP(1) receptor binding.     

The return of an experimentally N-saturated boreal forest to an N-limited state: observations on the soil microbial community structure,biotic N retention capacity and gross N mineralisation

Background and aims

To find out how N-saturated forests can return to an N-limited state, we examined the recovery of biotic N sinks under decreasing N supply.

Methods

. We studied a 40-year-old experiment in Pinus sylvestris forest, with control plots, N0, three N treatments, N1-N3, of which N3 was stopped after 20 years, allowing observation of recovery.

Results

In N3, the N concentration in foliage was still slightly elevated, but the N uptake capacity of ectomycorrhizal (ECM) roots in N3 was no longer lower than in N0. Per area the amount of a biomarker for fungi, here mainly attributed ECM, was higher in N3 and N0 than in N1 and N2. Retention of labeled ¹⁵NH₄ ⁺ by the soil was greater in the control (99 %) and N3 (86 %), than in N1 (45 %) and N2 (29 %); we ascribe these differences to biotic retention because cation exchange capacity did not vary. Gross N mineralisation and retention of N correlated, negatively and positively, respectively, with abundance of ECM fungal biomarker.

Conclusions

. The results suggest a key role for ECM fungi in regulating the N cycle. We propose, in accordance with plant C allocation theory, that recovery is driven by increased tree below-ground C allocation to ECM roots and fungi.     

Differences in late fetal death rates in association with determinants of small for gestational age fetuses: population based cohort study

Objective: To examine differences in late fetal death rates in association with determinants of small for gestational age fetuses. Design: Population based cohort study. Subjects: 1 026 249 pregnancies without congenital malformations. Setting: Sweden 1983-92. Main outcome measure: Late fetal death rate. Results: Depending on underlying determinants late fetal death rates were greatly increased in extremely small for gestational age fetuses (range 16 to 45 per 1000) compared with non-small for gestational age fetuses (1.4 to 4.6). In extremely small for gestational age fetuses late fetal death rates were increased from 31 per 1000 in mothers aged less than 35 years to 45 per 1000 in older mothers, and from 22 per 1000 in women <155 cm in height to 33 per 1000 in women ⩾175 cm tall. Late fetal death rates were also higher in extremely small for gestational age fetuses in singleton compared with twin pregnancies and in non-hypertensive pregnancies compared with pregnancies complicated by severe pre-eclampsia or other hypertensive disorders. Slightly higher late fetal death rates were observed in nulliparous compared with parous women and in non-smokers compared with smokers.Conclusions: Although the risk of late fetal death is greatly increased in fetuses that are extremely small for gestational age the risk is strongly modified by underlying determinants—for example, there is a lower risk of late fetal death in a small for gestational age fetus if the mother is of short stature, has a twin pregnancy, or has hypertension.

Key messages

• Small for gestational age fetuses are at increased risk of late fetal death regardless of the underlying determinants
• The effect of birthweight ratio on risk of late fetal death is modified by underlying determinants, except maternal age
• Regardless of birthweight ratio the rates of late fetal death are higher among women aged 35 years or older compared with younger women
• In pregnancies of extremely small for gestational age fetuses lower rates of late fetal death are associated with a maternal age of less than 35 years, short maternal stature, multiple births, and hypertensive disorders
• In pregnancies with non-malformed fetuses late fetal death rates are increased in smokers, in multiple births, and in women with severe pre-eclampsia.