Tweaking the NRF2 signaling cascade in human myelogenous leukemia cells by artificial nano-organelles.

NRF2 (nuclear factor erythroid-2-related factor 2) is a key regulator of genes involved in the cell's protective response to oxidative stress. Upon activation by disturbed redox homeostasis, NRF2 promotes the expression of metabolic enzymes to eliminate reactive oxygen species (ROS). Cell internalization of peroxisome-like artificial organelles that harbor redox-regulating enzymes was previously shown to reduce ROS-induced stress and thus cell death. However, if and to which extent ROS degradation by such nanocompartments interferes with redox signaling pathways is largely unknown. Here, we advance the design of H2O2-degrading artificial nano-organelles (AnOs) that exposed surface-attached cell penetrating peptides (CPP) for enhanced uptake and were equipped with a fluorescent moiety for rapid visualization within cells. To investigate how such AnOs integrate in cellular redox signaling, we engineered leukemic K562 cells that report on NRF2 activation by increased mCherry expression. Once internalized, ROS-metabolizing AnOs dampen intracellular NRF2 signaling upon oxidative injury by degrading H2O2. Moreover, intracellular AnOs conferred protection against ROSinduced cell death in conditions when endogenous ROS-protection mechanisms have been compromised by depletion of glutathione or knockdown of NRF2. We demonstrate CPP-facilitated AnO uptake and AnO-mediated protection against ROS insults also in the T lymphocyte population of primary peripheral blood mononuclear cells from healthy donors. Overall, our data suggest that intracellular AnOs alleviated cellular stress by the on-site reduction of ROS.

Advancing the Design of Artificial Nano-organelles for Targeted Cellular Detoxification of Reactive Oxygen Species.

Artificial organelles (AnOs) are in the spotlight as systems to supplement biochemical pathways in cells. While polymersome-based artificial organelles containing enzymes to reduce reactive oxygen species (ROS) are known, applications requiring control of their enzymatic activity and cell-targeting to promote intracellular ROS detoxification are underexplored. Here, we introduce advanced AnOs where the chemical composition of the membrane supports the insertion of pore-forming melittin, enabling molecular exchange between the AnO cavity and the environment, while the encapsulated lactoperoxidase (LPO) maintains its catalytic function. We show that H2O2 outside AnOs penetrates through the melittin pores and is rapidly degraded by the encapsulated enzyme. As surface attachment of cell-penetrating peptides facilitates AnOs uptake by cells, electron spin resonance revealed a remarkable enhancement in intracellular ROS detoxification by these cell-targeted AnOs compared to nontargeted AnOs, thereby opening new avenues for a significant reduction of oxidative stress in cells.

FAP Targeting of Photosensitizer-Loaded Polymersomes for Increased Light-Activated Cell Killing.

As current chemo- and photodynamic cancer therapies are associated with severe side effects due to a lack of specificity and to systemic toxicity, innovative solutions in terms of targeting and controlled functionality are in high demand. Here, we present the development of a polymersome nanocarrier equipped with targeting molecules and loaded with photosensitizers for efficient uptake and light-activated cell killing. Polymersomes were self-assembled in the presence of photosensitizers from a mixture of nonfunctionalized and functionalized PDMS-b-PMOXA diblock copolymers, the latter designed for coupling with targeting ligands. By encapsulation inside the polymersomes, the photosensitizer Rose Bengal was protected, and its uptake into cells was mediated by the nanocarrier. Inhibitor of fibroblast activation protein α (FAPi), a ligand for FAP, was attached to the polymersomes' surface and improved their uptake in MCF-7 breast cancer cells expressing relatively high levels of FAP on their surface. Once internalized by MCF-7, irradiation of Rose Bengal-loaded FAPi-polymersomes generated reactive oxygen species at levels high enough to induce cell death. By combining photosensitizer encapsulation and specific targeting, polymersomes represent ideal candidates as therapeutic nanocarriers in cancer treatment.

A DNA-Micropatterned Surface for Propagating Biomolecular Signals by Positional on-off Assembly of Catalytic Nanocompartments.

Signal transduction is pivotal for the transfer of information between and within living cells. The composition and spatial organization of specified compartments are key to propagating soluble signals. Here, a high-throughput platform mimicking multistep signal transduction which is based on a geometrically defined array of immobilized catalytic nanocompartments (CNCs) that consist of distinct polymeric nanoassemblies encapsulating enzymes and DNA or enzymes alone is presented. The dual role of single entities or tandem CNCs in providing confined but communicating spaces for complex metabolic reactions and in protecting encapsulated compounds from denaturation is explored. To support a controlled spatial organization of CNCs, CNCs are patterned by means of DNA hybridization to a microprinted glass surface. Specifically, CNC-functionalized DNA microarrays are produced where individual reaction compartments are kept in close proximity by a distinct geometrical arrangement to promote effective communication. Besides a remarkable versatility and robustness, the most prominent feature of this platform is the reversibility of DNA-mediated CNC-anchoring which renders it reusable. Micropatterns of polymer-based nanocompartment assemblies offer an ideal scaffold for the development of the next generation responsive and communicative soft-matter analytical devices for applications in catalysis and medicine.

Impact of a brochure and empathetic physician communication on patients' perception of breast biopsies.

PURPOSE: This study investigated the effect of an intervention designed to reduce patients' emotional distress associated with breast biopsy. METHODS: 125 breast biopsy patients receiving standard of care (control group, CG) were compared to 125 patients (intervention group, IG) who received a brochure with information prior to the biopsy and were biopsied by physicians trained in empathic communication. Anxiety was assessed by the State-Anxiety Inventory (STAI-S) at four time points (pre- and post-procedural, pre- and post-histology). All participants completed pre- and post-procedural questionnaires addressing worries, pain and comprehension. We evaluated the impact of the intervention on STAI-S levels using a log-transformed linear mixed effects model and explored patients' and physicians' perceptions of the procedure descriptively. RESULTS: Post-procedural and post-histology timepoints were associated with 13% and17% lower with STAI-S levels than at the pre-procedural timepoint on average. The histologic result had the strongest association with STAI-S: malignancy was associated with 28% higher STAI-S scores than a benign finding on average. Across all time points, the intervention did not affect patient anxiety. Nevertheless, IG participants perceived less pain during the biopsy. Nearly all patients agreed that the brochure should be handed out prior to breast biopsy. CONCLUSION: While the distribution of an informative brochure and a physician trained in empathic communication did not reduce patient anxiety overall, we observed lower levels of worry and perceived pain regarding breast biopsy in the intervention group. The intervention seemed to improve patient's understanding of the procedure. Moreover, professional training could increase physicians' empathic communication skills. TRIAL REGISTRATION NUMBER: NCT02796612 (March 19, 2014).

Agreement in breast lesion assessment and final BI-RADS classification between radial and meander-like breast ultrasound.

BACKGROUND: This study prospectively investigates the agreement between radial (r-US) and meander-like (m-US) breast ultrasound with regard to lesion location, lesion size, morphological characteristics and final BI-RADS classification of individual breast lesions. METHODS: Each patient of a consecutive, unselected, mixed collective received a dual ultrasound examination. RESULTS: The agreement between r-US and m-US for lesion location ranged from good (lesion to mammilla distance ICC 0.64; lesion to skin distance ICC 0.72) to substantial (clock-face localization κ 0.70). For lesion size the agreement was good (diameter ICC 0.72; volume ICC 0.69), for lesion margin and architectural distortion it was substantial (κ 0.68 and 0.70, respectively). Most importantly, there was a substantial agreement (κ 0.76) in the final BI-RADS classification between r-US and m-US. CONCLUSIONS: Our recent comparison of radial and meander-like breast US revealed that the diagnostic accuracy of the two scanning methods was comparable. In this study, we observe a high degree of agreement between m-US and r-US for the lesion description (location, size, morphology) and final BI-RADS classification. These findings corroborate that r-US is a suitable alternative to m-US in daily clinical practice. Trial registration NCT02358837. Registered January 2015, retrospectively registered https://clinicaltrials.gov/ct2/results?cond=&term=NCT02358837&cntry=&state=&city=&dist =.

Urotherapist activities in caring for patients with pelvic floor disorders: a prospective single-center observational study.

PURPOSE: Patients with pelvic floor disorders are growing in number. The aim of this study is to outline the main activities of a urotherapist, an advanced nurse practitioner, in the care of patients with pelvic floor disorders and to evaluate patient satisfaction with the service urotherapists provide. METHODS: The prospective single-center observational study was carried out from July 2016 to June 2018. Parameters used to assess the urotherapist activities included the number of consultations, type of counselling, time frame of consultations and therapy and patient satisfaction. In a subgroup of 38 patients, satisfaction with the urotherapy sessions was evaluated by a questionnaire. RESULTS: Totally, 1709 patients were examined by urogynecologists. Five hundred and fourteen (30%) with chronic pelvic floor disorders were subsequently referred to a urotherapist. Of these patients, 60% were at least 65 years old. The most common pelvic floor disorders (221 patients; 43%) were an overactive bladder, recurrent urinary tract infections, chronic cystitis and pelvic pain syndrome; the second most common pelvic floor disorder was pelvic organ prolapsed (151 patients; 29%). Of the study subgroup comprising 38 patients, 32 (84%) returned the patient satisfaction questionnaire. All 32 patients specified their level of agreement with the urotherapist's professional competence, empathy, temporal availability and quality of advice as "agree to strongly agree." CONCLUSIONS: Management by a urotherapist was highly appreciated. The role of the urotherapist as a care coordinator, their level of autonomy and barriers to the implementation in primary care requires further exploration.

Peptide-Assisted Nucleic Acid Delivery Systems on the Rise.

Concerns associated with nanocarriers' therapeutic efficacy and side effects have led to the development of strategies to advance them into targeted and responsive delivery systems. Owing to their bioactivity and biocompatibility, peptides play a key role in these strategies and, thus, have been extensively studied in nanomedicine. Peptide-based nanocarriers, in particular, have burgeoned with advances in purely peptidic structures and in combinations of peptides, both native and modified, with polymers, lipids, and inorganic nanoparticles. In this review, we summarize advances on peptides promoting gene delivery systems. The efficacy of nucleic acid therapies largely depends on cell internalization and the delivery to subcellular organelles. Hence, the review focuses on nanocarriers where peptides are pivotal in ferrying nucleic acids to their site of action, with a special emphasis on peptides that assist anionic, water-soluble nucleic acids in crossing the membrane barriers they encounter on their way to efficient function. In a second part, we address how peptides advance nanoassembly delivery tools, such that they navigate delivery barriers and release their nucleic acid cargo at specific sites in a controlled fashion.

DNA-tethered Polymersome Clusters as Nanotheranostic Platform.

Nanotheranostics combine the use of nanomaterials and biologically active compounds to achieve diagnosis and treatment at the same time. To date, severe limitations compromise the use of nanotheranostic systems as potent nanomaterials are often incompatible with potent biomolecules. Herein we emphasize how a novel type of polymersome clusters loaded with active molecules can be optimized to obtain an efficient nanotheranostic platform. Polymersomes loaded with enzymes and specific dyes, respectively and exposing complementary DNA strands at their external surface formed clusters by means of DNA hybridization. We describe factors at the molecular level and other conditions that need to be optimized at each step of the cluster formation to favor theranostic efficiency.

A self-assembling amphiphilic peptide nanoparticle for the efficient entrapment of DNA cargoes up to 100 nucleotides in length.

To overcome the low efficiency and cytotoxicity associated with most non-viral DNA delivery systems we developed a purely peptidic self-assembling system that is able to entrap single- and double-stranded DNA of up to 100 nucleotides in length. (HR)3gT peptide design consists of a hydrophilic domain prone to undergo electrostatic interactions with DNA cargo, and a hydrophobic domain at a ratio that promotes the self-assembly into multi-compartment micellar nanoparticles (MCM-NPs). Self-assembled (HR)3gT MCM-NPs range between 100 to 180 nm which is conducive to a rapid and efficient uptake by cells. (HR)3gT MCM-NPs had no adverse effects on HeLa cell viability. In addition, they exhibit long-term structural stability at 4 °C but at 37 °C, the multi-micellar organization disassembles overtime which demonstrates their thermo-responsiveness. The comparison of (HR)3gT to a shorter, less charged H3gT peptide indicates that the additional arginine residues result in the incorporation of longer DNA segments, an improved DNA entrapment efficiency and an increase cellular uptake. Our unique non-viral system for DNA delivery sets the stage for developing amphiphilic peptide nanoparticles as candidates for future systemic gene delivery.

Immobilization of arrestin-3 on different biosensor platforms for evaluating GPCR binding.

G protein-coupled receptors (GPCRs) are a large and ubiquitous family of membrane receptors of great pharmacological interest. Cell-based assays are the primary tool for assessing GPCR interactions and activation but their design and intrinsic complexity limit their application. Biosensor-based assays that directly and specifically report GPCR-protein binding (e.g. arrestin or G protein) could provide a good alternative. We present an approach based on the stable immobilization of different arrestin-3 proteins (wild type, and two mutants, mutant X (arrestin-3 I386A) and mutant Y (arrestin-3 R393E)) via histidine tags on NTA(Ni2+)-coated sensors in a defined orientation. Using biolayer interferometry (BLI), surface plasmon resonance (SPR), and quartz crystal microbalance with dissipation (QCM-D), we were able to follow the interaction between the different arrestin-3 proteins and a representative GPCR, jumping spider rhodopsin-1 (JSR1), in a label-free manner in real-time. The interactions were quantified as binding affinity, association and dissociation rate constants. The combination of surface-based biosensing methods indicated that JSR1 showed the strongest binding to arrestin mutant Y. Taken together, this work introduces direct label-free, biosensor-based screening approaches that can be easily adapted for testing interactions of proteins and other compounds with different GPCRs.

Comparison of radial and meander-like breast ultrasound with respect to diagnostic accuracy and examination time.

PURPOSE: To prospectively compare the diagnostic accuracy of radial breast ultrasound (r-US) to that of conventional meander-like breast ultrasound (m-US), patients of a consecutive, unselected, mixed collective were examined by both scanning methods. METHODS: Out of 1948 dual examinations, 150 revealed suspicious lesions resulting in 168 biopsies taken from 148 patients. Histology confirmed breast cancers in 36 cases. Sensitivity, specificity, accuracy, PPV, and NPV were calculated for r-US and m-US. The examination times were recorded. RESULTS: For m-US and r-US, sensitivity (both 88.9%), specificity (86.4% versus 89.4%), accuracy (86.9% versus 89.3%), PPV (64.0% versus 69.6%), NPV (both 98.3%), false-negative rate (both 5.6%), and rate of cancer missed by one method (both 5.6%) were similar. The mean examination time for r-US (14.8 min) was significantly (p < 0.01) shorter than for m-US (22.6 min). CONCLUSION: Because the diagnostic accuracy of r-US and m-US are comparable, r-US can be considered an alternative to m-US in routine breast US with the added benefit of a significantly shorter examination time.

Peptide-Based Nanoassemblies in Gene Therapy and Diagnosis: Paving the Way for Clinical Application.

Nanotechnology approaches play an important role in developing novel and efficient carriers for biomedical applications. Peptides are particularly appealing to generate such nanocarriers because they can be rationally designed to serve as building blocks for self-assembling nanoscale structures with great potential as therapeutic or diagnostic delivery vehicles. In this review, we describe peptide-based nanoassemblies and highlight features that make them particularly attractive for the delivery of nucleic acids to host cells or improve the specificity and sensitivity of probes in diagnostic imaging. We outline the current state in the design of peptides and peptide-conjugates and the paradigms of their self-assembly into well-defined nanostructures, as well as the co-assembly of nucleic acids to form less structured nanoparticles. Various recent examples of engineered peptides and peptide-conjugates promoting self-assembly and providing the structures with wanted functionalities are presented. The advantages of peptides are not only their biocompatibility and biodegradability, but the possibility of sheer limitless combinations and modifications of amino acid residues to induce the assembly of modular, multiplexed delivery systems. Moreover, functions that nature encoded in peptides, such as their ability to target molecular recognition sites, can be emulated repeatedly in nanoassemblies. Finally, we present recent examples where self-assembled peptide-based assemblies with "smart" activity are used in vivo. Gene delivery and diagnostic imaging in mouse tumor models exemplify the great potential of peptide nanoassemblies for future clinical applications.

Actin ADP-ribosylation at Threonine148 by Photorhabdus luminescens toxin TccC3 induces aggregation of intracellular F-actin.

Intoxication of eukaryotic cells by Photorhabdus luminescens toxin TccC3 induces cell rounding and detachment from the substratum within a few hours and compromises a number of cell functions like phagocytosis. Here, we used morphological and biochemical procedures to analyse the mechanism of TccC3 intoxication. Life imaging of TccC3-intoxicated HeLa cells transfected with AcGFP-actin shows condensation of F-actin into large aggregates. Life cell total internal reflection fluorescence (TIRF) microscopy of identically treated HeLa cells confirmed the formation of actin aggregates but also disassembly of F-actin stress fibres. Recombinant TccC3 toxin ADP-ribosylates purified skeletal and non-muscle actin at threonine148 leading to a strong propensity to polymerize and F-actin bundle formation as shown by TIRF and electron microscopy. Native gel electrophoresis shows strongly reduced binding of Thr148-ADP-ribosylated actin to the severing proteins gelsolin and its fragments G1 and G1-3, and to ADF/cofilin. Complexation of actin with these proteins inhibits its ADP-ribosylation. TIRF microscopy demonstrates rapid polymerization of Thr148-ADP-ribosylated actin to curled F-actin bundles even in the presence of thymosin ß4, gelsolin or G1-3. Thr148-ADP-ribosylated F-actin cannot be depolymerized by gelsolin or G1-3 as verified by TIRF, co-sedimentation and electron microscopy and shows reduced treadmilling as indicated by a lack of stimulation of its ATPase activity after addition of cofilin-1.

Biopores/membrane proteins in synthetic polymer membranes.

BACKGROUND: Mimicking cell membranes by simple models based on the reconstitution of membrane proteins in lipid bilayers represents a straightforward approach to understand biological function of these proteins. This biomimetic strategy has been extended to synthetic membranes that have advantages in terms of chemical and mechanical stability, thus providing more robust hybrid membranes. SCOPE OF THE REVIEW: We present here how membrane proteins and biopores have been inserted both in the membrane of nanosized and microsized compartments, and in planar membranes under various conditions. Such bio-hybrid membranes have new properties (as for example, permeability to ions/molecules), and functionality depending on the specificity of the inserted biomolecules. Interestingly, membrane proteins can be functionally inserted in synthetic membranes provided these have appropriate properties to overcome the high hydrophobic mismatch between the size of the biomolecule and the membrane thickness. MAJOR CONCLUSION: Functional insertion of membrane proteins and biopores in synthetic membranes of compartments or in planar membranes is possible by an appropriate selection of the amphiphilic copolymers, and conditions of the self-assembly process. These hybrid membranes have new properties and functionality based on the specificity of the biomolecules and the nature of the synthetic membranes. GENERAL SIGNIFICANCE: Bio-hybrid membranes represent new solutions for the development of nanoreactors, artificial organelles or active surfaces/membranes that, by further gaining in complexity and functionality, will promote translational applications. This article is part of a Special Issue entitled: Lipid order/lipid defects and lipid-control of protein activity edited by Dirk Schneider.

Contributions of the lower dimer to supramolecular actin patterning revealed by TIRF microscopy.

Two distinct dimers are formed during the initial steps of actin polymerization. The first one, referred to as the 'lower dimer' (LD) was discovered many years ago by means of chemical crosslinking. Owing to its transient nature, a biological relevance had long been precluded when, using LD-specific antibodies, we detected LD-like contacts in actin assemblies that are associated with the endolysosomal compartment in a number of different cell lines. Moreover, immunofluorescence showed the presence of LD-related structures at the cell periphery of migrating fibroblasts, in the nucleus, and in association with the centrosome of interphase cells. Here, we explore contributions of the LD to the assembly of supramolecular actin structures in real time by total internal reflection fluorescence (TIRF) microscopy. Our data shows that while LD on its own cannot polymerize under filament forming conditions, it is able to incorporate into growing F-actin filaments. This incorporation of LD triggers the formation of X-shaped filament assemblies with barbed ends that are pointing in the same direction in the majority of cases. Similarly, an increased frequency of junction sites was observed when filaments were assembled in the presence of oxidized actin. This data suggests that a disulfide bridge between Cys374 residues might stabilize LD-contacts. Based on our findings, we propose two possible models for the molecular mechanism underlying the supramolecular actin patterning in LD-related structures.

FHOD1 is a combined actin filament capping and bundling factor that selectively associates with actin arcs and stress fibers.

Formins are actin polymerization factors that are known to nucleate and elongate actin filaments at the barbed end. In the present study we show that human FHOD1 lacks actin nucleation and elongation capacity, but acts as an actin bundling factor with capping activity toward the filament barbed end. Constitutively active FHOD1 associates with actin filaments in filopodia and lamellipodia at the leading edge, where it moves with the actin retrograde flow. At the base of lamellipodia, FHOD1 is enriched in nascent, bundled actin arcs as well as in more mature stress fibers. This function requires actin-binding domains located N-terminally to the canonical FH1-FH2 element. The bundling phenotype is maintained in the presence of tropomyosin, confirmed by electron microscopy showing assembly of 5 to 10 actin filaments into parallel, closely spaced filament bundles. Taken together, our data suggest a model in which FHOD1 stabilizes actin filaments by protecting barbed ends from depolymerization with its dimeric FH2 domain, whereas the region N-terminal to the FH1 domain mediates F-actin bundling by simultaneously binding to the sides of adjacent F-actin filaments.

Synthetic Cells Revisited: Artificial Cells Construction Using Polymeric Building Blocks.

The exponential growth of research on artificial cells and organelles underscores their potential as tools to advance the understanding of fundamental biological processes. The bottom-up construction from a variety of building blocks at the micro- and nanoscale, in combination with biomolecules is key to developing artificial cells. In this review, artificial cells are focused upon based on compartments where polymers are the main constituent of the assembly. Polymers are of particular interest due to their incredible chemical variety and the advantage of tuning the properties and functionality of their assemblies. First, the architectures of micro- and nanoscale polymer assemblies are introduced and then their usage as building blocks is elaborated upon. Different membrane-bound and membrane-less compartments and supramolecular structures and how they combine into advanced synthetic cells are presented. Then, the functional aspects are explored, addressing how artificial organelles in giant compartments mimic cellular processes. Finally, how artificial cells communicate with their surrounding and each other such as to adapt to an ever-changing environment and achieve collective behavior as a steppingstone toward artificial tissues, is taken a look at. Engineering artificial cells with highly controllable and programmable features open new avenues for the development of sophisticated multifunctional systems.

Nanoassemblies designed for efficient nuclear targeting.

One of the key aspects of coping efficiently with complex pathological conditions is delivering the desired therapeutic compounds with precision in both space and time. Therefore, the focus on nuclear-targeted delivery systems has emerged as a promising strategy with high potential, particularly in gene therapy and cancer treatment. Here, we explore the design of supramolecular nanoassemblies as vehicles to deliver specific compounds to the nucleus, with the special focus on polymer and peptide-based carriers that expose nuclear localization signals. Such nanoassemblies aim at maximizing the concentration of genetic and therapeutic agents within the nucleus, thereby optimizing treatment outcomes while minimizing off-target effects. A complex scenario of conditions, including cellular uptake, endosomal escape, and nuclear translocation, requires fine tuning of the nanocarriers' properties. First, we introduce the principles of nuclear import and the role of nuclear pore complexes that reveal strategies for targeting nanosystems to the nucleus. Then, we provide an overview of cargoes that rely on nuclear localization for optimal activity as their integrity and accumulation are crucial parameters to consider when designing a suitable delivery system. Considering that they are in their early stages of research, we present various cargo-loaded peptide- and polymer nanoassemblies that promote nuclear targeting, emphasizing their potential to enhance therapeutic response. Finally, we briefly discuss further advancements for more precise and effective nuclear delivery.

Patient perception of meander-like versus radial breast ultrasound.

Background Radial breast ultrasound scanning (r-US) and commonly used meander-like ultrasound scanning (m-US) have recently been shown to be equally sensitive and specific with regard to the detection of breast malignancies. As patient satisfaction has a strong influence on patient compliance and thus on the quality of health care, we compare here the two US scanning techniques with regard to patient comfort during breast ultrasound (BUS) and analyze whether the patient has a preference for either scanning technique. Materials and Methods Symptomatic and asymptomatic women underwent both m-US and r-US scanning by two different examiners. Patient comfort and preference were assessed using a visual analog scale-based (VAS) questionnaire and were compared using a Mann-Whitney U test. Results Analysis of 422 VAS-based questionnaires showed that perceived comfort with r-US (r-VAS 8 cm, IQR [5.3, 9.1]) was significantly higher compared to m-US (m-VAS 5.6 cm, IQR [5.2, 7.4]) (p < 0.001). 53.8% of patients had no preference, 44.3% of patients clearly preferred r-US, whereas only 1.9% of patients preferred m-US. Conclusion: Patients experience a higher level of comfort with r-US and favor r-US over m-US. As the diagnostic accuracy of r-US has been shown to be comparable to that of m-US and the time required for examination is shorter, a switch from m-US to r-US in routine clinical practice might be beneficial. R-US offers considerable potential to positively affect patient compliance but also to save examination time and thus costs.