Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) and Endolysosomal Two-pore Channels Modulate Membrane Excitability and Stimulus-Secretion Coupling in Mouse Pancreatic ß Cells.

Pancreatic ß cells are electrically excitable and respond to elevated glucose concentrations with bursts of Ca(2+) action potentials due to the activation of voltage-dependent Ca(2+) channels (VDCCs), which leads to the exocytosis of insulin granules. We have examined the possible role of nicotinic acid adenine dinucleotide phosphate (NAADP)-mediated Ca(2+) release from intracellular stores during stimulus-secretion coupling in primary mouse pancreatic ß cells. NAADP-regulated Ca(2+) release channels, likely two-pore channels (TPCs), have recently been shown to be a major mechanism for mobilizing Ca(2+) from the endolysosomal system, resulting in localized Ca(2+) signals. We show here that NAADP-mediated Ca(2+) release from endolysosomal Ca(2+) stores activates inward membrane currents and depolarizes the ß cell to the threshold for VDCC activation and thereby contributes to glucose-evoked depolarization of the membrane potential during stimulus-response coupling. Selective pharmacological inhibition of NAADP-evoked Ca(2+) release or genetic ablation of endolysosomal TPC1 or TPC2 channels attenuates glucose- and sulfonylurea-induced membrane currents, depolarization, cytoplasmic Ca(2+) signals, and insulin secretion. Our findings implicate NAADP-evoked Ca(2+) release from acidic Ca(2+) storage organelles in stimulus-secretion coupling in ß cells.

Incidence and outcomes of vasa praevia in the United Kingdom.

Background: Vasa praevia is an obstetric condition in which the fetal vessels run through the membrane over the internal cervical os, unprotected by the placenta or umbilical cord. It is associated with perinatal mortality if not diagnosed antenatally. We investigated the incidence and outcomes of vasa praevia in the UK. Methods: We conducted a population-based descriptive study using the UK Obstetric Surveillance System (UKOSS). Cases were identified prospectively through monthly UKOSS submissions form all UK hospitals with obstetrician-led maternity units. All women diagnosed with vasa praevia who gave birth between 1 st December 2014 and 30 th November 2015 were included. The main outcome was incidence of vasa praevia with 95% confidence intervals, using 2015 maternities as the denominator. Results: Fifty-one women met the case definition. The incidence of diagnosed vasa praevia was 6.64 per 100,000 maternities (95% CI 5.05-8.73). Of 198 units, 10 (5%) had a vasa praevia screening programme; one of these 10 units identified 25% of the antenatally diagnosed cases. Among women who had vasa praevia diagnosed or suspected antenatally (n=28, 55%), there were no perinatal deaths or hypoxic ischaemic encephalopathy (HIE). Twenty-four women with antenatal diagnosis were hospitalised at a median gestation of 32 weeks and caesarean section was scheduled at a median gestation of 36 weeks. When vasa praevia was diagnosed peripartum (n=23, 45%), the perinatal mortality rate was 37.5% and 47% of survivors developed HIE. Conclusions: The incidence of diagnosed vasa praevia was lower than anticipated. There was high perinatal mortality and morbidity for cases not diagnosed antenatally. The incidence of antenatally identified cases was much higher in the few centres that actively screened for this condition, and the perinatal outcomes were better. However, this group were all delivered by caesarean section and may include women who would not have experienced any adverse perinatal outcome.

Vasa praevia is a pregnancy complication in which the blood vessels that connect the mother and fetus run across the opening of the womb, without protection from the placenta or umbilical cord. During birth, the vessels can tear. This can result in rapid blood loss from the baby and in some cases, death of the baby. We investigated how common vasa praevia is in the UK, and how women with the condition and their babies fared. The UK Obstetric Surveillance System (UKOSS) collects anonymous information from all maternity units in the UK about pregnant women who have certain medical conditions. UKOSS reporters provided information about all women with vasa praevia who gave birth between December 2014 and November 2015. We identified 51 women with vasa praevia, meaning vasa praevia was diagnosed less often in the UK than we had expected based on studies from other countries. Twenty-eight women were diagnosed during the antenatal period, while 23 were diagnosed during labour or after giving birth. Pregnant women in the UK are not screened for vasa praevia as standard, and some women may have had vasa praevia that was not diagnosed. A small number (5%) of maternity units in our study did offer screening for vasa praevia in their pregnant population. One of these units identified a quarter of all the women who had vasa praevia diagnosed during pregnancy. Babies born to women whose vasa praevia was diagnosed during pregnancy had good outcomes. All of these women gave birth by planned caesarean section, and they and their babies survived. Babies born to women whose vasa praevia was suspected or diagnosed during labour or after birth had worse outcomes. Around 40% were stillborn or died shortly after birth, and about half of those who survived had brain damage caused by lack of oxygen.

Methods used to develop the SPIRIT 2024 and CONSORT 2024 Statements.

OBJECTIVES: To describe, and explain the rationale for, the methods used and decisions made during development of the updated SPIRIT 2024 and CONSORT 2024 reporting guidelines. METHODS: We developed SPIRIT 2024 and CONSORT 2024 together to facilitate harmonization of the two guidelines, and incorporated content from key extensions. We conducted a scoping review of comments suggesting changes to SPIRIT 2013 and CONSORT 2010, and compiled a list of other possible revisions based on existing SPIRIT and CONSORT extensions, other reporting guidelines, and personal communications. From this, we generated a list of potential modifications or additions to SPIRIT and CONSORT, which we presented to stakeholders for feedback in an international online Delphi survey. The Delphi survey results were discussed at an online expert consensus meeting attended by 30 invited international participants. We then drafted the updated SPIRIT and CONSORT checklists and revised them based on further feedback from meeting attendees. RESULTS: We compiled 83 suggestions for revisions or additions to SPIRIT and/or CONSORT from the scoping review and 85 from other sources, from which we generated 33 potential changes to SPIRIT (n = 5) or CONSORT (n = 28). Of 463 participants invited to take part in the Delphi survey, 317 (68%) responded to Round 1, 303 (65%) to Round 2 and 290 (63%) to Round 3. Two additional potential checklist changes were added to the Delphi survey based on Round 1 comments. Overall, 14/35 (SPIRIT n = 0; CONSORT n = 14) proposed changes reached the predefined consensus threshold (≥80% agreement), and participants provided 3580 free-text comments. The consensus meeting participants agreed with implementing 11/14 of the proposed changes that reached consensus in the Delphi and supported implementing a further 4/21 changes (SPIRIT n = 2; CONSORT n = 2) that had not reached the Delphi threshold. They also recommended further changes to refine key concepts and for clarity. CONCLUSION: The forthcoming SPIRIT 2024 and CONSORT 2024 Statements will provide updated, harmonized guidance for reporting randomized controlled trial protocols and results, respectively. The simultaneous development of the SPIRIT and CONSORT checklists has been informed by current empirical evidence and extensive input from stakeholders. We hope that this report of the methods used will be helpful for developers of future reporting guidelines.

Absence of intracellular ion channels TPC1 and TPC2 leads to mature-onset obesity in male mice, due to impaired lipid availability for thermogenesis in brown adipose tissue.

Intracellular calcium-permeable channels have been implicated in thermogenic function of murine brown and brite/beige adipocytes, respectively transient receptor potential melastin-8 and transient receptor potential vanilloid-4. Because the endo-lysosomal two-pore channels (TPCs) have also been ascribed with metabolic functionality, we studied the effect of simultaneously knocking out TPC1 and TPC2 on body composition and energy balance in male mice fed a chow diet. Compared with wild-type mice, TPC1 and TPC2 double knockout (Tpcn1/2(-/-)) animals had a higher respiratory quotient and became obese between 6 and 9 months of age. Although food intake was unaltered, interscapular brown adipose tissue (BAT) maximal temperature and lean-mass adjusted oxygen consumption were lower in Tpcn1/2(-/-) than in wild type mice. Phosphorylated hormone-sensitive lipase expression, lipid density and expression of ß-adrenergic receptors were also lower in Tpcn1/2(-/-) BAT, whereas mitochondrial respiratory chain function and uncoupling protein-1 expression remained intact. We conclude that Tpcn1/2(-/-) mice show mature-onset obesity due to reduced lipid availability and use, and a defect in ß-adrenergic receptor signaling, leading to impaired thermogenic activity, in BAT.

TPC1 has two variant isoforms, and their removal has different effects on endo-lysosomal functions compared to loss of TPC2.

Organelle ion homeostasis within the endo-lysosomal system is critical for physiological functions. Two-pore channels (TPCs) are cation channels that reside in endo-lysosomal organelles, and overexpression results in endo-lysosomal trafficking defects. However, the impact of a lack of TPC expression on endo-lysosomal trafficking is unknown. Here, we characterize Tpcn1 expression in two transgenic mouse lines (Tpcn1(XG716) and Tpcn1(T159)) and show expression of a novel evolutionarily conserved Tpcn1B transcript from an alternative promoter, raising important questions regarding the status of Tpcn1 expression in mice recently described to be Tpcn1 knockouts. We show that the transgenic Tpcn1(T159) line lacks expression of both Tpcn1 isoforms in all tissues analyzed. Using mouse embryonic fibroblasts (MEFs) from Tpcn1(-/-) and Tpcn2(-/-) animals, we show that a lack of Tpcn1 or Tpcn2 expression has no significant impact on resting endo-lysosomal pH or morphology. However, differential effects in endo-lysosomal function were observed upon the loss of Tpcn1 or Tpcn2 expression; thus, while Tpcn1(-/-) MEFs have impaired trafficking of cholera toxin from the plasma membrane to the Golgi apparatus, Tpcn2(-/-) MEFs show slower kinetics of ligand-induced platelet-derived growth factor receptor ß (PDGFRß) degradation, which is dependent on trafficking to lysosomes. Our findings indicate that TPC1 and TPC2 have important but distinct roles in the endo-lysosomal pathway.