Enhanced Ca2+ signaling, mild primary aldosteronism, and hypertension in a familial hyperaldosteronism mouse model (Cacna1hM1560V/+ ).

Gain-of-function mutations in the CACNA1H gene (encoding the T-type calcium channel CaV3.2) cause autosomal-dominant familial hyperaldosteronism type IV (FH-IV) and early-onset hypertension in humans. We used CRISPR/Cas9 to generate Cacna1hM1560V/+ knockin mice as a model of the most common FH-IV mutation, along with corresponding knockout mice (Cacna1h-/- ). Adrenal morphology of both Cacna1hM1560V/+ and Cacna1h-/- mice was normal. Cacna1hM1560V/+ mice had elevated aldosterone:renin ratios (a screening parameter for primary aldosteronism). Their adrenal Cyp11b2 (aldosterone synthase) expression was increased and remained elevated on a high-salt diet (relative autonomy, characteristic of primary aldosteronism), but plasma aldosterone was only elevated in male animals. The systolic blood pressure of Cacna1hM1560V/+ mice was 8 mmHg higher than in wild-type littermates and remained elevated on a high-salt diet. Cacna1h-/- mice had elevated renal Ren1 (renin-1) expression but normal adrenal Cyp11b2 levels, suggesting that in the absence of CaV3.2, stimulation of the renin-angiotensin system activates alternative calcium entry pathways to maintain normal aldosterone production. On a cellular level, Cacna1hM1560V/+ adrenal slices showed increased baseline and peak intracellular calcium concentrations in the zona glomerulosa compared to controls, but the frequency of calcium spikes did not rise. We conclude that FH-IV, on a molecular level, is caused by elevated intracellular Ca2+ concentrations as a signal for aldosterone production in adrenal glomerulosa cells. We demonstrate that a germline Cacna1h gain-of-function mutation is sufficient to cause mild primary aldosteronism, whereas loss of CaV3.2 channel function can be compensated for in a chronic setting.

Functions of the (pro)renin receptor (Atp6ap2) at molecular and system levels: pathological implications in hypertension, renal and brain development, inflammation, and fibrosis.

The (pro)renin receptor [(P)RR, Atp6ap2] was initially discovered as a membrane-bound binding partner of prorenin and renin. A soluble (P)RR has additional paracrine effects and is involved in metabolic syndrome and kidney damage. Meanwhile it is clear that most of the effects of the (P)RR are independent of prorenin. In the kidney, (P)RR plays an important role in renal dysfunction by activating proinflammatory and profibrotic molecules. In the brain, (P)RR is expressed in cardiovascular regulatory nuclei and is linked to hypertension. (P)RR is known to be an essential component of the v-ATPase as a key accessory protein and plays an important role in kidney, brain and heart via regulating the pH of the extracellular space and intracellular compartments. V-ATPase and (P)RR together act on WNT and mTOR signalling pathways, which are responsible for cellular homeostasis and autophagy. (P)RR through its role in v-ATPase assembly and function is also important for fast recycling endocytosis by megalin. In the kidney, megalin together with v-ATPase and (P)RR is crucial for endocytic uptake of components of the RAS and their intracellular processing. In the brain, (P)RR, v-ATPases and megalin are important regulators both during development and in the adult. All three proteins are associated with diseases such as XLMR, XMRE, X-linked parkinsonism and epilepsy, cognitive disorders with Parkinsonism, spasticity, intellectual disability, and Alzheimer's Disease which are characterized by impaired neuronal function and/or neuronal loss. The present review focusses on the relevant effects of Atp6ap2 without assigning them necessarily to the RAS. Mechanistically, many effects can be well explained by the role of Atp6ap2 for v-ATPase assembly and function. Furthermore, application of a soluble (P)RR analogue as new therapeutic option is discussed.

Refinable tri-variate C 1 splines for box-complexes including irregular points and irregular edges.

C 1 splines over box-complexes generalize C 1 degree 3 (cubic) tensor-product splines. A box-complex is a collection of 3-dimensional boxes forming an unstructured hexahedral mesh that can include irregular points and irregular edges where the layout deviates from the tensor-product grid layout. For example, an edge shared and enclosed by five boxes is irregular. Where the mesh is locally regular, the restriction of the space to each box is a polynomial piece of the C 1 tri-cubic tensor-product spline, by default initialized as a C 2 tri-cubic. Boxes containing irregularities have their polynomials binarily split into 23 pieces to isolate the irregularity. The pieces join with matching derivatives. The derivatives are zero at irregularities, but these singularities are removable by a local change of variables. The space consists of 23 linearly independent functions per box and is refinable.

Low degree splines for locally quad-dominant meshes.

A mesh is locally quad-dominant (lqd) if all non-4-sided facets are surrounded by quadrilaterals. Lqd meshes allow for irregular nodes where n ≠ 4 quads meet and for multi-sided facets, called T-gons, that end quad-strips and so adjust mesh density. This paper introduces a new class of bi-cubic (bi-3) Geometric T-joint (GT) splines whose control nets are τ-nets, i.e. T-gons surrounded by quads. These GT-splines join smoothly with each other, bi-3 G-splines and regular C 1 bi-quadratic splines to form smooth surfaces of degree at most bi-3.

Smooth polar caps for locally quad-dominant meshes.

A polar configuration is a node surrounded by m triangles. Polar configurations are common to cap off cylinders and spheres. When the triangles, interpreted as quadrilaterals with one edge collapsed, are surrounded by a quad-strip then the extended polar configuration qualifies as part of a locally quad-dominant (lqd) mesh. Recent constructions, referred to as semi-structured splines, can use lqd meshes as control nets: multi-sided configurations that merge parameter directions are covered by G-spline; and T-junctions that transition from coarse and fine are covered by GT-splines. This paper complements existing semi-structured splines by providing the missing component for polar configurations. A spectrum of constructions of differing degree are introduced, tested and compared. Bi-2 C 1 splines are extended to polar configurations covered by C 1 surfaces consisting of (macro-)patches of degree as low as bi-2. Bi-3 C 2 splines are extended to polar configurations covered by surfaces that are C 2 except for a C 1 pole and consist of (macro-)patches of degree as low as bi-3.

Localized G-splines for quad & T-gon meshes.

Enriching tensor-product B-spline control nets by allowing T-gons (where strips of quadrilaterals start or end) and irregular nodes (where n ≠ 4 quadrilaterals meet) reduces the requirements on quad-meshing and increases the flexibility for polyhedral design with associated smooth surfaces. This paper introduces a family of piecewise polynomial, geometrically continuous surface constructions that yield good highlight line distributions also in the presence of irregular nodes next to a T-gon. Such tight juxtaposition can further reduce the quad-meshing requirements and increase the space of polyhedral design control structures. The surfaces can be chosen to cover T-gons with G 1 caps of degree bi-4 - or with caps of degree bi-3 that are almost G 1 and preserve the good highlight line distribution of the bi-4 G 1 surfaces.

Fair free-form surfaces that are almost everywhere parametrically C 2.

Compared to G k continuity, C k continuity simplifies the construction of functions on surfaces and their refinement, e.g. to solve differential equations on the surface. The new class of almost everywhere parametrically C 2 free-form surfaces provide such a parameterization. For example, a new bi-6 construction combines a fast-contracting C 2 guided subdivision surface with a tiny multi-sided G 1 cap. The cap is chosen to be smaller than any refinement anticipated for geometric modeling or computing on surfaces. Fast contraction means that one subdivision step shrinks the remaining hole more than three steps of Catmull-Clark subdivision. This yields smooth surfaces consisting of a finite number of pieces that are suitable for engineering practice. Both the subdivision construction and the cap are guided by a reference surface. This guide conveys the basic shape, but has a different structure and lower smoothness.

Refinable smooth surfaces for locally quad-dominant meshes with T-gons.

Multi-sided facets in polyhedral models and meshes serve to connect regular submeshes (star-configurations) and to start or end quad-strips (T-configurations). Using the polyhedral mesh as control net, recursive subdivision algorithms often yield poor shape for these non-quad configurations. Polynomial surface constructions such as geometrically smooth splines (G-splines) do better, but lack subdivision-like refinability. Such refinability is useful for hierarchical modeling and engineering analysis. This paper introduces a new class of G-splines that generalizes bi-quadratic C 1 splines to polyhedral control nets with star- and T-configurations and that is refinable.

An alternative renin isoform is cardioprotective by modulating mitochondrial metabolism.

The renin-angiotensin system promotes oxidative stress, apoptosis, necrosis, fibrosis, and thus heart failure. Secretory renin plays a central role in these processes, initiating the generation of angiotensins. Nevertheless, alternative renin transcripts exist, which code for a cytosolically localized renin isoform (cyto-renin) that is cardioprotective. We tested the hypothesis that the protective effects are associated with a beneficial switch of metabolic and mitochondrial functions. To assess H9c2 cell mitochondrial parameters, we used the Seahorse XF analyser. Cardiac H9c2 cells overexpressing cyto-renin exhibited enhanced nonmitochondrial oxygen consumption, lactate accumulation, and LDH activity, reflecting a switch to more aerobic glycolysis known as Warburg effect. Additionally, mitochondrial spare capacity and cell respiratory control ratio were enhanced, indicating an increased potential to tolerate stress conditions. Renin knockdown induced opposite effects on mitochondrial functions without influencing metabolic parameters. Thus, the protective effects of cyto-renin are associated with an altered bioenergetic profile and an enhanced stress tolerance, which are favourable under ischaemic conditions. Therefore, cyto-renin is a promising new target for the prevention of ischaemia-induced myocardial damage.

The angiotensin II type 2 receptors protect renal tubule mitochondria in early stages of diabetes mellitus.

Diabetic nephropathy correlates more closely to defective mitochondria and increased oxidative stress in the kidney than to hyperglycemia. A key driving factor of diabetic nephropathy is angiotensin II acting via the G-protein-coupled cell membrane type 1 receptor. The present study aimed to investigate the role of the angiotensin II type 2 receptor (AT2R) at the early stages of diabetic nephropathy. Using receptor binding studies and immunohistochemistry we found that the mitochondria in renal tubules contain high-affinity AT2Rs. Increased renal mitochondrial AT2R density by transgenic overexpression was associated with reduced superoxide production of isolated mitochondria from non-diabetic rats. Streptozotocin-induced diabetes (28 days) caused a drop in the ATP/oxygen ratio and an increase in the superoxide production of isolated renal mitochondria from wild-type diabetic rats. This correlated with changes in the renal expression profile and increased tubular epithelial cell proliferation. AT2R overexpression in tubular epithelial cells inhibited all diabetes-induced renal changes including a drop in mitochondrial bioenergetics efficiency, a rise in mitochondrial superoxide production, metabolic reprogramming, and increased proliferation. Thus, AT2Rs translocate to mitochondria and can contribute to reno-protective effects at early stages of diabetes. Hence, targeted AT2R overexpression in renal cells may open new avenues to develop novel types of drugs preventing diabetic nephropathy.

(Pro)renin receptor (ATP6AP2) depletion arrests As4.1 cells in the G0/G1 phase thereby increasing formation of primary cilia.

The (pro)renin receptor [(P)RR, ATP6AP2] is a multifunctional transmembrane protein that activates local renin-angiotensin systems, but also interacts with Wnt pathways and vacuolar H+ -ATPase (V-ATPase) during organogenesis. The aim of this study was to characterize the role of ATP6AP2 in the cell cycle in more detail. ATP6AP2 down-regulation by siRNA in renal As4.1 cells resulted in a reduction in the rate of proliferation and a G0/G1 phase cell cycle arrest. We identified a number of novel target genes downstream of ATP6AP2 knock-down that were related to the primary cilium (Bbs-1, Bbs-3, Bbs-7, Rabl5, Ttc26, Mks-11, Mks-5, Mks-2, Tctn2, Nme7) and the cell cycle (Pierce1, Clock, Ppif). Accordingly, the number of cells expressing the primary cilium was markedly increased. We found no indication that these effects were dependent of V-ATPase activity, as ATP6AP2 knock-down did not affect lysosomal pH and bafilomycin A neither influenced the ciliary expression pattern nor the percentage of ciliated cells. Furthermore, ATP6AP2 appears to be essential for mitosis. ATP6AP2 translocated from the endoplasmatic reticulum to mitotic spindle poles (pro-, meta- and anaphase) and the central spindle bundle (telophase) and ATP6AP2 knock-down results in markedly deformed spindles. We conclude that ATP6AP2 is necessary for cell division, cell cycle progression and mitosis. ATP6AP2 also inhibits ciliogenesis, thus promoting proliferation and preventing differentiation.

The (pro)renin receptor and its interaction partners.

The prorenin receptor was originally discovered as a receptor that binds renin and prorenin, thereby inducing pro-fibrotic intracellular signal cascades. These effects are partially mediated in vitro by angiotensin (ANG) and partially independent of ANG. Consequently, inhibitors of the interaction between the prorenin and the (pro)renin receptor (PRR) were designed hoping that they may prevent fibrotic tissue damage, for instance, in the kidney. However, this concept was challenged by the fact that overexpression of the PRR was not harmful at all, whereas depletion of the PRR was lethal or markedly detrimental. Furthermore, the high levels of prorenin needed to activate the PRR may not be reached in vivo. As it turned out, the PRR instead exhibited a variety of important functions that has nothing to do with the name given to the protein. Thus, the PRR was identified as an accessory subunit of vesicular (v)-ATPases, representing an essential chaperon for the assembly of v-ATPase subunits. In this respect, the gene encoding the PRR was also named ATP6AP2. Finally, the PRR is an essential component of the canonical and non-canonical PCP Wnt pathways. Thus, the PRR is essential for lysosomal functions, such as endocytosis, secretion, and autophagy as well as for cell division and differentiation, embryonic development, organogenesis, and stem cell biology.

An Alternative Promoter in Intron1 of the Renin Gene is Regulated by Glucose Starvation via Serum Response Factor.

BACKGROUND/AIMS: Renin is known as a secretory glycoprotein that ultimately leads to angiotensin II generation. In this way renin exerts pro-inflammatory effects and promotes cardiac injury. Additional transcripts have been identified encoding for a cytosolic renin isoform that - in contrast to secretory renin - exhibits cardioprotective effects under ischemic conditions. The promoter of these transcripts is unknown. METHODS: Using qRT-PCR and dual-luciferase reporter assay we examined the expression and promotor activity of cytosolic renin as well as the regulation by glucose starvation in H9c2 cardiomyoblasts. RESULTS: We identified a promoter in intron1 of the rat renin gene with two glucose starvation-sensitive regions. One region contains a binding motif for serum response factor (SRF). Under glucose depletion expression of SRF increased prior to cytosolic renin. SRF knock down selectively decreased cytosolic renin expression and attenuated the increase of cytosolic renin expression under glucose depletion. CONCLUSIONS: Transcripts encoding for secretory and cytosolic renin are differentially expressed. The low basal expression of cytosolic renin as well as its induction under ischemia-related conditions represents an efficient system regulated in accordance with its previously identified unfavorable effects under control situations but protective effects seen after myocardial infarction or glucose depletion.

Cross-linguistic vowel variation in trilingual speakers of Saterland Frisian, Low German, and High German.

The present study compares the acoustic realization of Saterland Frisian, Low German, and High German vowels by trilingual speakers in the Saterland. The Saterland is a rural municipality in northwestern Germany. It offers the unique opportunity to study trilingualism with languages that differ both by their vowel inventories and by external factors, such as their social status and the autonomy of their speech communities. The objective of the study was to examine whether the trilingual speakers differ in their acoustic realizations of vowel categories shared by the three languages and whether those differences can be interpreted as effects of either the differences in the vowel systems or of external factors. Monophthongs produced in a /hVt/ frame revealed that High German vowels show the most divergent realizations in terms of vowel duration and formant frequencies, whereas Saterland Frisian and Low German vowels show small differences. These findings suggest that vowels of different languages are likely to share the same phonological space when the speech communities largely overlap, as is the case with Saterland Frisian and Low German, but may resist convergence if at least one language is shared with a larger, monolingual speech community, as is the case with High German.

A cross-dialectal acoustic study of Saterland Frisian vowels.

Previous investigations on Saterland Frisian report a large vowel inventory, including up to 20 monophthongs and 16 diphthongs in stressed position. Conducting a cross-dialectal acoustic study on Saterland Frisian vowels in Ramsloh, Scharrel, and Strücklingen, the objective is to provide a phonetic description of vowel category realization and to identify acoustic dimensions which may enhance the discrimination of neighboring categories within the crowded vowel space of the endangered minority language. All vowels were elicited in a /hVt/ frame. Acoustic measurements included vowel duration, mid-vowel F1 and F2, and the amount of spectral change and the spectral rate of change. The results suggest instances of phonetic attrition, i.e., merged categories, precisely where contrasts were reported to be on the retreat. The cross-dialectal comparison showed differences between the three dialects primarily within the F1 dimension at the 20% measurement point. The findings presented here contribute to the description of an endangered minority language and add to the question of which acoustic variables are used in languages with large vowel inventories to maintain or enhance the existing contrasts. Furthermore, the results underline the importance of the consideration of inter-speaker variability as well as measurements beyond the vowel target.

Surgeon-Authored Virtual Laparoscopic Adrenalectomy Module Is Judged Effective and Preferred Over Traditional Teaching Tools.

OBJECTIVE: The study assesses user acceptance and effectiveness of a surgeon-authored virtual reality (VR) training module authored by surgeons using the Toolkit for Illustration of Procedures in Surgery (TIPS). METHODS: Laparoscopic adrenalectomy was selected to test the TIPS framework on an unusual and complex procedure. No commercial simulation module exists to teach this procedure. A specialist surgeon authored the module, including force-feedback interactive simulation, and designed a quiz to test knowledge of the key procedural steps. Five practicing surgeons, with 15 to 24 years of experience, peer reviewed and tested the module. In all, 14 residents and 9 fellows trained with the module and answered the quiz, preuse and postuse. Participants received an overview during Surgical Grand Rounds session and a 20-minute one-on-one tutorial followed by 30 minutes of instruction in addition to a force-feedback interactive simulation session. Additionally, in answering questionnaires, the trainees reflected on their learning experience and their experience with the TIPS framework. RESULTS: Correct quiz response rates on procedural steps improved significantly postuse over preuse. In the questionnaire, 96% of the respondents stated that the TIPS module prepares them well or very well for the adrenalectomy, and 87% indicated that the module successfully teaches the steps of the procedure. All participants indicated that they preferred the module compared to training using purely physical props, one-on-one teaching, medical atlases, and video recordings. CONCLUSIONS: Improved quiz scores and endorsement by the participants of the TIPS adrenalectomy module establish the viability of surgeons authoring VR training.

The Wnt adaptor protein ATP6AP2 regulates multiple stages of adult hippocampal neurogenesis.

In the mammalian hippocampus, canonical Wnt signals provided by the microenvironment regulate the differentiation of adult neural stem cells (NSCs) toward the neuronal lineage. Wnts are part of a complex and diverse set of signaling pathways and the role of Wnt/Planar cell polarity (PCP) signaling in adult neurogenesis remains unknown. Using in vitro assays on differentiating adult NSCs, we identified a transition of Wnt signaling responsiveness from Wnt/ß-catenin to Wnt/PCP signaling. In mice, retroviral knockdown strategies against ATP6AP2, a recently discovered core protein involved in both signaling pathways, revealed that its dual role is critical for granule cell fate and morphogenesis. We were able to confirm its dual role in neurogenic Wnt signaling in vitro for both canonical Wnt signaling in proliferating adult NSCs and non-canonical Wnt signaling in differentiating neuroblasts. Although LRP6 appeared to be critical for granule cell fate determination, in vivo knockdown of PCP core proteins FZD3 and CELSR1-3 revealed severe maturational defects without changing the identity of newborn granule cells. Furthermore, we found that CELSR1-3 control distinctive aspects of PCP-mediated granule cell morphogenesis with CELSR1 regulating the direction of dendrite initiation sites and CELSR2/3 controlling radial migration and dendritic patterning. The data presented here characterize distinctive roles for Wnt/ß-catenin signaling in granule cell fate determination and for Wnt/PCP signaling in controlling the morphological maturation of differentiating neuroblasts.

Refinable C1 spline elements for irregular quad layout.

Building on a result of U. Reif on removable singularities, we construct C1 bi-3 splines that may include irregular points where less or more than four tensor-product patches meet. The resulting space complements PHT splines, is refinable and the refined spaces are nested, preserving for example surfaces constructed from the splines. As in the regular case, each quadrilateral has four degrees of freedom, each associated with one spline and the splines are linearly independent. Examples of use for surface construction and isogeometric analysis are provided.

C1 finite elements on non-tensor-product 2d and 3d manifolds.

Geometrically continuous (Gk ) constructions naturally yield families of finite elements for isogeometric analysis (IGA) that are Ck also for non-tensor-product layout. This paper describes and analyzes one such concrete C1 geometrically generalized IGA element (short: gIGA element) that generalizes bi-quadratic splines to quad meshes with irregularities. The new gIGA element is based on a recently-developed G1 surface construction that recommends itself by its a B-spline-like control net, low (least) polynomial degree, good shape properties and reproduction of quadratics at irregular (extraordinary) points. Remarkably, for Poisson's equation on the disk using interior vertices of valence 3 and symmetric layout, we observe O(h3) convergence in the L∞ norm for this family of elements. Numerical experiments confirm the elements to be effective for solving the trivariate Poisson equation on the solid cylinder, deformations thereof (a turbine blade), modeling and computing geodesics on smooth free-form surfaces via the heat equation, for solving the biharmonic equation on the disk and for Koiter-type thin-shell analysis.