Structural basis for inhibition of the lysosomal two-pore channel TPC2 by a small molecule antagonist.

Two pore channels are lysosomal cation channels with crucial roles in tumor angiogenesis and viral release from endosomes. Inhibition of the two-pore channel 2 (TPC2) has emerged as potential therapeutic strategy for the treatment of cancers and viral infections, including Ebola and COVID-19. Here, we demonstrate that antagonist SG-094, a synthetic analog of the Chinese alkaloid medicine tetrandrine with increased potency and reduced toxicity, induces asymmetrical structural changes leading to a single binding pocket at only one intersubunit interface within the asymmetrical dimer. Supported by functional characterization of mutants by Ca2+ imaging and patch clamp experiments, we identify key residues in S1 and S4 involved in compound binding to the voltage sensing domain II. SG-094 arrests IIS4 in a downward shifted state which prevents pore opening via the IIS4/S5 linker, hence resembling gating modifiers of canonical VGICs. These findings may guide the rational development of new therapeutics antagonizing TPC2 activity.

In-house 3D-printed custom splints for non-operative treatment of distal radial fractures: a randomized controlled trial.

We compared patient satisfaction and clinical effectiveness of 3D-printed splints made of photopolymer resin to conventional fibre glass casts in treating distal radial fractures. A total of 39 patients with minimally displaced distal radius fractures were included and randomized. Of them, 20 were immobilized in a fibre glass cast and 19 in a 3D-printed forearm splint. The 3D-printed splints were custom-designed based on forearm surface scanning with a handheld device and printed in-house using digital light processing printing technology. Patient satisfaction and clinical effectiveness were assessed with questionnaires 1 and 6 weeks after the initiation of immobilization. Fracture healing, pain, range of motion, grip strength and the DASH and PRWE scores were assessed up to 1-year follow-up. 3D-printed splints proved to be equally well tolerated by the patients and equally clinically effective as conventional fibre glass casts although there was a higher rate of minor complications. 3D-printed splints present a safe alternative, especially in young, active patients, for non-operative treatment of distal radial fractures.Level of evidence: I.

Two-pore channel-2 and inositol trisphosphate receptors coordinate Ca2+ signals between lysosomes and the endoplasmic reticulum.

Lysosomes and the endoplasmic reticulum (ER) are Ca2+ stores mobilized by the second messengers NAADP and IP3, respectively. Here, we establish Ca2+ signals between the two sources as fundamental building blocks that couple local release to global changes in Ca2+. Cell-wide Ca2+ signals evoked by activation of endogenous NAADP-sensitive channels on lysosomes comprise both local and global components and exhibit a major dependence on ER Ca2+ despite their lysosomal origin. Knockout of ER IP3 receptor channels delays these signals, whereas expression of lysosomal TPC2 channels accelerates them. High-resolution Ca2+ imaging reveals elementary events upon TPC2 opening and signals coupled to IP3 receptors. Biasing TPC2 activation to a Ca2+-permeable state sensitizes local Ca2+ signals to IP3. This increases the potency of a physiological agonist to evoke global Ca2+ signals and activate a downstream target. Our data provide a conceptual framework to understand how Ca2+ release from physically separated stores is coordinated.

Convergent activation of Ca2+ permeability in two-pore channel 2 through distinct molecular routes.

TPC2 is a pathophysiologically relevant lysosomal ion channel that is activated directly by the phosphoinositide PI(3,5)P2 and indirectly by the calcium ion (Ca2+)-mobilizing molecule NAADP through accessory proteins that associate with the channel. TPC2 toggles between PI(3,5)P2-induced, sodium ion (Na+)-selective and NAADP-induced, Ca2+-permeable states in response to these cues. To address the molecular basis of polymodal gating and ion-selectivity switching, we investigated the mechanism by which NAADP and its synthetic functional agonist, TPC2-A1-N, induced Ca2+ release through TPC2 in human cells. Whereas NAADP required the NAADP-binding proteins JPT2 and LSM12 to evoke endogenous calcium ion signals, TPC2-A1-N did not. Residues in TPC2 that bind to PI(3,5)P2 were required for channel activation by NAADP but not for activation by TPC2-A1-N. The cryptic voltage-sensing region of TPC2 was required for the actions of TPC2-A1-N and PI(3,5)P2 but not for those of NAADP. These data mechanistically distinguish natural and synthetic agonist action at TPC2 despite convergent effects on Ca2+ permeability and delineate a route for pharmacologically correcting impaired NAADP-evoked Ca2+ signals.

Extension osteotomy of the metacarpal I and ligamentoplasty of the trapeziometacarpal joint for the treatment of early-stage osteoarthritis and instability of the trapeziometacarpal joint.

Osteoarthritis (OA) is a common disease of the first carpo-metacarpal (CMC I) joint. Biomechanical factors promoting OA are the shape of the CMC I-joint, being a biconcave-convex saddle joint with high mobility and the increased instability caused by joint space narrowing, ligamentous laxity, and direction of force transmission of the abductor pollicis longus (APL) tendon during adduction. The closing wedge osteotomy of the base of the first metacarpal is joint preserving treatment option. We combine this closing wedge osteotomy with a ligamentoplasty to stabilize the joint. In this manuscript, we provide a detailed description of the indication, discuss biomechanical aspects and the surgical technique in detail.

Artificial intelligence in patient-specific hand surgery: a scoping review of literature.

PURPOSE: The implementation of artificial intelligence in hand surgery and rehabilitation is gaining popularity. The purpose of this scoping review was to give an overview of implementations of artificial intelligence in hand surgery and rehabilitation and their current significance in clinical practice. METHODS: A systematic literature search of the MEDLINE/PubMed and Cochrane Collaboration libraries was conducted. The review was conducted according to the framework outlined by the Preferred Reporting Items for Systematic Reviews and Meta-Analysis Extension for Scoping Reviews. A narrative summary of the papers is presented to give an orienting overview of this rapidly evolving topic. RESULTS: Primary search yielded 435 articles. After application of the inclusion/exclusion criteria and addition of supplementary search, 235 articles were included in the final review. In order to facilitate navigation through this heterogenous field, the articles were clustered into four groups of thematically related publications. The most common applications of artificial intelligence in hand surgery and rehabilitation target automated image analysis of anatomic structures, fracture detection and localization and automated screening for other hand and wrist pathologies such as carpal tunnel syndrome, rheumatoid arthritis or osteoporosis. Compared to other medical subspecialties the number of applications in hand surgery is still small. CONCLUSION: Although various promising applications of artificial intelligence in hand surgery and rehabilitation show strong performances, their implementation mostly takes place within the context of experimental studies. Therefore, their use in daily clinical routine is still limited.

Expanding the Toolbox: Novel Modulators of Endolysosomal Cation Channels.

Functional characterization of endolysosomal ion channels is challenging due to their intracellular location. With recent advances in endolysosomal patch clamp technology, it has become possible to directly measure ion channel currents across endolysosomal membranes. Members of the transient receptor potential (TRP) cation channel family, namely the endolysosomal TRPML channels (TRPML1-3), also called mucolipins, as well as the distantly related two-pore channels (TPCs) have recently been characterized in more detail with endolysosomal patch clamp techniques. However, answers to many physiological questions require work in intact cells or animal models. One major obstacle thereby is that the known endogenous ligands of TRPMLs and TPCs are anionic in nature and thus impermeable for cell membranes. Microinjection, on the other hand, is technically demanding. There is also a risk of losing essential co-factors for channel activation or inhibition in isolated preparations. Therefore, lipophilic, membrane-permeable small-molecule activators and inhibitors for TRPMLs and TPCs are urgently needed. Here, we describe and discuss the currently available small-molecule modulators of TRPMLs and TPCs.

Distance mapping in three-dimensional virtual surgical planning in hand, wrist and forearm surgery: a tool to avoid mistakes.

PURPOSE: Three-dimensional planning in corrective surgeries in the hand and wrist has become popular throughout the last 20 years. Imaging technologies and software have improved since their first description in the late 1980s. New imaging technologies, such as distance mapping (DM), improve the safety of virtual surgical planning (VSP) and help to avoid mistakes. We describe the effective use of DM in two representative and frequently performed surgical interventions (radius malunion and scaphoid pseudoarthrosis). METHODS: We simulated surgical intervention in both cases using DM. Joint spaces were quantitatively and qualitatively displayed in a colour-coded fashion, which allowed the estimation of cartilage thickness and joint space congruency. These parameters are presented in the virtual surgical planning pre- and postoperatively as well as in the actual situation in our cases. RESULTS: DM had a high impact on the VSP, especially in radius corrective osteotomy, where we changed the surgical plan due to the visualization of the planned postoperative situation. The actual postoperative situation was also documented using DM, which allowed for comparison of the VSP and the achieved postoperative situation. Both patients were successfully treated, and bone healing and clinical improvement were achieved. CONCLUSION: The use of colour-coded static or dynamic distance mapping is useful for virtual surgical planning of corrective osteotomies of the hand, wrist and forearm. It also allows confirmation of the correct patient treatment and assessment of the follow-up radiological documentation.

TPC2 rescues lysosomal storage in mucolipidosis type IV, Niemann-Pick type C1, and Batten disease.

Lysosomes are cell organelles that degrade macromolecules to recycle their components. If lysosomal degradative function is impaired, e.g., due to mutations in lysosomal enzymes or membrane proteins, lysosomal storage diseases (LSDs) can develop. LSDs manifest often with neurodegenerative symptoms, typically starting in early childhood, and going along with a strongly reduced life expectancy and quality of life. We show here that small molecule activation of the Ca2+ -permeable endolysosomal two-pore channel 2 (TPC2) results in an amelioration of cellular phenotypes associated with LSDs such as cholesterol or lipofuscin accumulation, or the formation of abnormal vacuoles seen by electron microscopy. Rescue effects by TPC2 activation, which promotes lysosomal exocytosis and autophagy, were assessed in mucolipidosis type IV (MLIV), Niemann-Pick type C1, and Batten disease patient fibroblasts, and in neurons derived from newly generated isogenic human iPSC models for MLIV and Batten disease. For in vivo proof of concept, we tested TPC2 activation in the MLIV mouse model. In sum, our data suggest that TPC2 is a promising target for the treatment of different types of LSDs, both in vitro and in-vivo.

Segregated cation flux by TPC2 biases Ca2+ signaling through lysosomes.

Two-pore channels are endo-lysosomal cation channels with malleable selectivity filters that drive endocytic ion flux and membrane traffic. Here we show that TPC2 can differentially regulate its cation permeability when co-activated by its endogenous ligands, NAADP and PI(3,5)P2. Whereas NAADP rendered the channel Ca2+-permeable and PI(3,5)P2 rendered the channel Na+-selective, a combination of the two increased Ca2+ but not Na+ flux. Mechanistically, this was due to an increase in Ca2+ permeability independent of changes in ion selectivity. Functionally, we show that cell permeable NAADP and PI(3,5)P2 mimetics synergistically activate native TPC2 channels in live cells, globalizing cytosolic Ca2+ signals and regulating lysosomal pH and motility. Our data reveal that flux of different ions through the same pore can be independently controlled and identify TPC2 as a likely coincidence detector that optimizes lysosomal Ca2+ signaling.

The ethoxycarbonyl group as both activating and protective group in N-acyl-Pictet-Spengler reactions using methoxystyrenes. A short approach to racemic 1-benzyltetrahydroisoquinoline alkaloids.

We present a systematic investigation on an improved variant of the N-acyl-Pictet-Spengler condensation for the synthesis of 1-benzyltetrahydroisoquinolines, based on our recently published synthesis of N-methylcoclaurine, exemplified by the total syntheses of 10 alkaloids in racemic form. Major advantages are a) using ω-methoxystyrenes as convenient alternatives to arylacetaldehydes, and b) using the ethoxycarbonyl residue for both activating the arylethylamine precursors for the cyclization reaction, and, as a significant extension, also as protective group for phenolic residues. After ring closure, the ethoxycarbonyl-protected phenols are deprotected simultaneously with the further processing of the carbamate group, either following route A (lithium alanate reduction) to give N-methylated phenolic products, or following route B (treatment with excess methyllithium) to give the corresponding alkaloids with free N-H function. This dual use of the ethoxycarbonyl group shortens the synthetic routes to hydroxylated 1-benzyltetrahydroisoquinolines significantly. Not surprisingly, these ten alkaloids did not show noteworthy effects on TPC2 cation channels and the tumor cell line VCR-R CEM, and did not exhibit P-glycoprotein blocking activity. But due to their free phenolic groups they can serve as valuable intermediates for novel derivatives addressing all of these targets, based on previous evidence for structure-activity relationships in this chemotype.

Overview of In-Hospital 3D Printing and Practical Applications in Hand Surgery.

Three-dimensional (3D) printing is spreading in hand surgery. There is an increasing number of practical applications like the training of junior hand surgeons, patient education, preoperative planning, and 3D printing of customized casts, customized surgical guides, implants, and prostheses. Some high-quality studies highlight the value for surgeons, but there is still a lack of high-level evidence for improved clinical endpoints and hence actual impact on the patient's outcome. This article provides an overview over the latest applications of 3D printing in hand surgery and practical experience of implementing them into daily clinical routine.

A Rare Case of Cutaneous Trichophyton verrucosum of the Forearm in a 51-Year-Old Cattle Farmer.

Trichophyton verrucosum is a pathogen causing superficial mycoses in cattle worldwide and is one of the few zoophilic dermatophytes. Farmers and veterinarians are at a higher risk for infection owing to frequent direct animal contact. An increase in cases among humans has been observed in the past few years. We report a rare case of T verrucosum of the forearm in a 51-year-old cattle farmer, who after initial treatment with antibiotics and surgery, and in whom diagnosis was delayed, was finally successfully treated with terbinafine and itraconazole.

[Implant arthroplasty in the metacarpophalangeal and proximal interphalangeal finger joints]. / Übersicht über die Endoprothetik an den Fingergrund- und Mittelgelenken.

BACKGROUND: Osteoarthritic changes in the finger joints are common, especially in the elderly population. Without adequate treatment, these changes can lead to pain, joint deformity, instability or impaired motion. Operative treatment options can be divided into prosthetic joint replacement, joint fusion and denervation. PATIENTS/MATERIAL AND METHOD: During the last decades, various prosthetic implant designs have appeared on the market. This article provides an overview of implant evolution, current implants, clinical results and promising technical novelties. RESULTS: Due to favourable clinical long-term results, low revision rates and low costs, the proven silicone spacer has been the gold standard since the 1960 s. In the index and middle finger, lateral stability is crucial to providing a counter bearing to the thumb for a strong key pinch. Medullary-anchored prostheses and modular surface replacement designs have a higher intrinsic stability and may thus be advantageous in the index and middle finger. These implants show promising clinical medium-term results. CONCLUSION: In the past, technical novelties from big joint replacements could not automatically be translated to the finger joints and other parts of the hand. However, new trends such as customised or 3D-printed prosthetic implants are slowly beginning to gain importance in hand surgery.

Chemical and pharmacological characterization of the TRPML calcium channel blockers ML-SI1 and ML-SI3.

The members of the TRPML subfamily of non-selective cation channels (TRPML1-3) are involved in the regulation of important lysosomal and endosomal functions, and mutations in TRPML1 are associated with the neurodegenerative lysosomal storage disorder mucolipidosis type IV. For in-depth investigation of functions and (patho)physiological roles of TRPMLs, membrane-permeable chemical tools are urgently needed. But hitherto only two TRPML inhibitors, ML-SI1 and ML-SI3, have been published, albeit without clear information about stereochemical details. In this investigation we developed total syntheses of both inhibitors. ML-SI1 was only obtained as a racemic mixture of inseparable diastereomers and showed activator-dependent inhibitory activity. The more promising tool is ML-SI3, hence ML-SI1 was not further investigated. For ML-SI3 we confirmed by stereoselective synthesis that the trans-isomer is significantly more active than the cis-isomer. Separation of the enantiomers of trans-ML-SI3 further revealed that the (-)-isomer is a potent inhibitor of TRPML1 and TRPML2 (IC50 values 1.6 and 2.3 µM) and a weak inhibitor (IC50 12.5 µM) of TRPML3, whereas the (+)-enantiomer is an inhibitor on TRPML1 (IC50 5.9 µM), but an activator on TRPML 2 and 3. This renders the pure (-)-trans-ML-SI3 more suitable as a chemical tool for the investigation of TRPML1 and 2 than the racemate. The analysis of 12 analogues of ML-SI3 gave first insights into structure-activity relationships in this chemotype, and showed that a broad variety of modifications in both the N-arylpiperazine and the sulfonamide moiety is tolerated. An aromatic analogue of ML-SI3 showed an interesting alternative selectivity profile (strong inhibitor of TRPML1 and strong activator of TRPML2).

[Implant Arthroplasty in the Hand and Wrist]. / Endoprothetik an der Hand und am Handgelenk.

Implant Arthroplasty in the Hand and Wrist Abstract. The first implant arthroplasties were conducted as early as the end of the 19th century. After a few decades, during which surgeons focused on the hip and knee, research on joint arthroplasty of the hand and wrist was intensified. Today, it is possible to implant a prosthesis in nearly every joint of the hand and wrist, giving us an alternative to joint fusion in severe joint destruction. The main advantage of arthroplasty is the preservation of motion. In the metacarpophalangeal and proximal interphalangeal joint every new implant is rivaled by the silicone spacer which has proven its value over decades. Further, no implant in the thumb carpometacarpal joint was able to outmatch the time-tested trapezectomy with LRTI (ligament reconstruction and tendon interposition) due to high implant-loosening rates in the past. In wrist arthroplasty, studies have shown good results concerning pain reduction, function and grip strength. Complication rates however are still higher compared to joint fusion. The results improved with the latest implant designs, yet long-term outcomes, especially for the young, high-demand patients, are missing.

Clinical and radiological results of the vascularized medial femoral condyle graft for scaphoid non-union.

INTRODUCTION: This study evaluated the use of a free vascularized bone graft with and without cartilage from the medial femoral condyle (MFC) in patients with recalcitrant scaphoid non-union, with a special focus on union rates and the osteochondral graft for proximal pole destruction. MATERIALS AND METHODS: Thirty-eight avascular scaphoid non-unions in 37 patients who were treated with a free osteoperiosteal or osteochondral MFC graft were retrospectively evaluated (mean follow-up 16 months). Bone union, the scapholunate and the radiolunate angles were evaluated on X-ray images. The range of motion, grip strength, VAS, DASH and PRWE scores were evaluated clinically. RESULTS: The overall union rate was 95%. Bone union was achieved in 27 out of 29 (93%) scaphoids treated with a free osteoperiosteal MFC grafts and in 9 out of 9 (100%) scaphoids treated with a free osteochondral MFC graft. The range of motion remained almost unchanged, while grip strength increased significantly (34 kg vs. 44 kg) and the VAS (22-5), DASH (59-19) and PRWE (62-30) score decreased significantly. The scapholunate (71°-65°) and radiolunate (28°-18°) angle decreased. No major donor site morbidity was observed. Postoperative complications were observed in eight cases (21%). CONCLUSIONS: The vascularized medial femoral bone graft leads to a good functional outcome in the treatment of scaphoid non-unions. The graft provides adequate blood supply and structural stability to the scaphoid. A proximal pole destruction can be replaced using an osteochondral graft with promising short-term results preventing carpal osteoarthritis and collapse.

Agonist-mediated switching of ion selectivity in TPC2 differentially promotes lysosomal function.

Ion selectivity is a defining feature of a given ion channel and is considered immutable. Here we show that ion selectivity of the lysosomal ion channel TPC2, which is hotly debated (Calcraft et al., 2009; Guo et al., 2017; Jha et al., 2014; Ruas et al., 2015; Wang et al., 2012), depends on the activating ligand. A high-throughput screen identified two structurally distinct TPC2 agonists. One of these evoked robust Ca2+-signals and non-selective cation currents, the other weaker Ca2+-signals and Na+-selective currents. These properties were mirrored by the Ca2+-mobilizing messenger, NAADP and the phosphoinositide, PI(3,5)P2, respectively. Agonist action was differentially inhibited by mutation of a single TPC2 residue and coupled to opposing changes in lysosomal pH and exocytosis. Our findings resolve conflicting reports on the permeability and gating properties of TPC2 and they establish a new paradigm whereby a single ion channel mediates distinct, functionally-relevant ionic signatures on demand.

Hook Plate for Volar Rim Fractures of the Distal Radius: Review of the First 23 Cases and Focus on Dorsal Radiocarpal Dislocation.

Background Fragment-specific fixation of the distal radius is born to fix each articular fragment with limited surgical approach and low-profile devices. Over time, many devices with different designs and characteristics have been developed. However, many of them have showed the inability to securely fix marginal, small, and comminuted fragments as bony ligament avulsions and bony compression injuries. Purpose The purpose of this study was to evaluate the clinical and radiological outcome of a new device born to treat marginal articular fractures of the distal radius. Patients and Methods A retrospective review was conducted on 23 patients with a mean follow-up of 21 months including postoperative clinical evaluation, grip strength, computed tomography scan, and X-ray control. Results All fragments healed and maintained reduced until the final follow-up. The carpus was aligned with the distal radius in all patients presenting with a radiocarpal dislocation. Conclusion The volar rim fragment is an attachment site for the short radiolunate and the volar distal radioulnar ligament. Its unstable fixation can lead to articular incongruity, volar or dorsal subluxation of the carpus, and distal radioulnar instability. The involvement of this fragment on distal radius fractures is relatively common and many studies of the literature have been focused on its treatment. The Hook Plate stabilizes distal fragments at their bone-ligament interface. In addition to bony reduction, the device permits to stabilize the capsule and ligaments, as volar bony ligament avulsions, in a picture of dorsal radiocarpal dislocation. Level of Evidence This is a Level IV, case series.