Liquid-liquid phase separation underpins the formation of replication factories in rotaviruses.

RNA viruses induce the formation of subcellular organelles that provide microenvironments conducive to their replication. Here we show that replication factories of rotaviruses represent protein-RNA condensates that are formed via liquid-liquid phase separation of the viroplasm-forming proteins NSP5 and rotavirus RNA chaperone NSP2. Upon mixing, these proteins readily form condensates at physiologically relevant low micromolar concentrations achieved in the cytoplasm of virus-infected cells. Early infection stage condensates could be reversibly dissolved by 1,6-hexanediol, as well as propylene glycol that released rotavirus transcripts from these condensates. During the early stages of infection, propylene glycol treatments reduced viral replication and phosphorylation of the condensate-forming protein NSP5. During late infection, these condensates exhibited altered material properties and became resistant to propylene glycol, coinciding with hyperphosphorylation of NSP5. Some aspects of the assembly of cytoplasmic rotavirus replication factories mirror the formation of other ribonucleoprotein granules. Such viral RNA-rich condensates that support replication of multi-segmented genomes represent an attractive target for developing novel therapeutic approaches.

The indication of fusion to the pelvis in neuromuscular scoliosis is based on the underlying disease rather than on pelvic obliquity.

INTRODUCTION: The decision to instrument to L5 or ilium, in NMS, is usually based on radiologic factors, including pelvic obliquity (PO) > 15°, apex of curvature < L3, and Cobb angle > 60°. Since scoliosis in these patients is caused by a neurologic disease, we based our decision to stop at L5 on the presence of spasticity or flaccidity. PATIENTS & METHODS: The senior author did 109 primary fusions in NMS. Of those with DMD or SMA only 16% were instrumented to the ilium. The main factor for our decision was the correction potential of the truncal shift and PO in the supine traction radiographs and the absence of severe spasticity. RESULTS: The 57 patients with DMD/SMA had a mean preoperative curvature of 68°, PO of 17°, and truncal shift of 20°. 74% should have been instrumented to the pelvis, but only 16% were. Those instrumented shorter as the rule, were corrected from 74° to 26° and had a postoperative PO of 8°. There was no significant difference in postoperative correction and PO compared to those instrumented to L5 on standard protocol. Subsequent extension to the pelvis was needed in 1 CP patient. There were no significant changes after 2 years. Of the 20 patients instrumented to the pelvis 11 had cerebral palsy and a preop curvature of 89°, a PO of 21° and a truncal shift of 25°. DISCUSSION: The decision on instrumentation length should take flexibility and disease into consideration. If the trunk is centred over the pelvis, deterioration will not occur in absence of spasticity.

Derotation screws provide no advantage over polyaxial screws regarding coronal & sagittal correction in thoracic curves of AIS patients.

PURPOSE: We compared two techniques for thoracic apical derotation; one using conventional reduction screws (Single-Innie-SI) and one requiring special derotation screws that can be converted to monoaxial screws to enhance dorotation (Dual-Innie-DI) for coronal and sagittal correction and. METHODS: A total of 200 patients with thoracic AIS have been included. In the SI-Group (n = 127) the convex rod was applied first. Vertebral derotation was done by translation to the concave rod with the convex rod being in place and center of rotation (COR). In the DI-Group (n = 73) correction started with translation on the concave side as well but now followed by derotation around the concave rod using the DI-mechanism. RESULTS: The mean rotation according to Raimondi and coronal correction was not sig. affected (72 (± 12) % in the SI-Group versus 68 (± 15) % in the DI-Group), even when flexibility was respected (Cincinnati Correction Index CCI was 2.9 (± 4.9) versus 3.5 (± 4.4). (p < 0.01). The gain of kyphosis was sig greater (2.7°) in the SI-group, but not clinical relevant. CONCLUSION: The use of DI screws for apical derotation did not provide an advantage for coronal correction or derotation in thoracic curves. Presumably after translation is performed in the DI-group, there was too much tension and friction in the construct impeding further derotation. Simultaneous translation and derotation in the SI-group, with the convex rod being the COR, yielded similar correction with better kyphosis and was faster and more economic.

Artificial intelligence facilitates decision-making in the treatment of lumbar disc herniations.

PURPOSE: Apart from patients with severe neurological deficits, it is not clear whether surgical or conservative treatment of lumbar disc herniations is superior for the individual patient. We investigated whether deep learning techniques can predict the outcome of patients with lumbar disc herniation after 6 months of treatment. METHODS: The data of 60 patients were used to train and test a deep learning algorithm with the aim to achieve an accurate prediction of the ODI 6 months after surgery or the start of conservative therapy. We developed an algorithm that predicts the ODI of 6 randomly selected test patients in tenfold cross-validation. RESULTS: A 100% accurate prediction of an ODI range could be achieved by dividing the ODI scale into 12% sections. A maximum absolute difference of only 3.4% between individually predicted and actual ODI after 6 months of a given therapy was achieved with our most powerful model. The application of artificial intelligence as shown in this work also allowed to compare the actual patient values after 6 months with the prediction for the alternative therapy, showing deviations up to 18.8%. CONCLUSION: Deep learning in the supervised form applied here can identify patients at an early stage who would benefit from conservative therapy, and on the contrary avoid painful and unnecessary delays for patients who would profit from surgical therapy. In addition, this approach can be used in many other areas of medicine as an effective tool for decision-making when choosing between opposing treatment options, despite small patient groups.

Supersensitive Multifluorophore RNA-FISH for Early Virus Detection and Flow-FISH by Using Click Chemistry.

The reliable detection of transcription events through the quantification of the corresponding mRNA is of paramount importance for the diagnostics of infections and diseases. The quantification and localization analysis of the transcripts of a particular gene allows disease states to be characterized more directly compared to an analysis on the transcriptome wide level. This is particularly needed for the early detection of virus infections as now required for emergent viral diseases, e. g. Covid-19. In situ mRNA analysis, however, is a formidable challenge and currently performed with sets of single-fluorophore-containing oligonucleotide probes that hybridize to the mRNA in question. Often a large number of probe strands (>30) are required to get a reliable signal. The more oligonucleotide probes are used, however, the higher the potential off-target binding effects that create background noise. Here, we used click chemistry and alkyne-modified DNA oligonucleotides to prepare multiple-fluorophore-containing probes. We found that these multiple-dye probes allow reliable detection and direct visualization of mRNA with only a very small number (5-10) of probe strands. The new method enabled the in situ detection of viral transcripts as early as 4 hours after infection.

Dendrimer-Based Signal Amplification of Click-Labelled DNA in Situ.

The in vivo incorporation of alkyne-modified bases into the genome of cells is today the basis for the efficient detection of cell proliferation. Cells are grown in the presence of ethinyl-dU (EdU), fixed and permeabilised. The incorporated alkynes are then efficiently detected by using azide-containing fluorophores and the CuI -catalysed alkyne-azide click reaction. For a world in which constant improvement in the sensitivity of a given method is driving diagnostic advancement, we developed azide- and alkyne-modified dendrimers that allow the establishment of sandwich-type detection assays that show significantly improved signal intensities and signal-to-noise ratios far beyond that which is currently possible.

Is the anterior approach still superior to posterior correction in AIS regarding correction, fusion levels and kyphosis when modern posterior systems are used?

PURPOSE: The aim of our study is to compare anterior and posterior corrections of thoracic (Lenke I) and lumbar (Lenke V) curves when modern posterior pedicle screw systems with vertebral derotation techniques are used. Curves that could not be corrected with both systems were excluded. METHODS: A thoracic group (N = 56) of Lenke I AIS patients (18 anterior and 38 posterior) and a lumbar group (N = 42) of Lenke V patients (14 anterior and 28 posterior) with similar curves < 65° were identified. RESULTS: Thoracic group The mean postoperative correction (POC) was 68 ± 13.4% in the anterior and 72 ± 10.5% in the posterior group. The postoperative change in thoracic kyphosis was +4° and +5° respectively. The median length of fusion was eight segments in the posterior and seven segments in the anterior groups. In 89% the LIV was EV or shorter in the anterior, and in 71% of the posterior corrections. Lumbar group The mean POC was 75 ± 18.3% (anterior) and 72 ± 8.5% (posterior). The postoperative gain in lumbar lordosis was 0.8° (anterior) and 4° (posterior). The median length of fusion was five segments in both groups and there was no difference in relation of the LIV to the EV. CONCLUSION: With modern implants and derotation techniques, the posterior approach can achieve similar coronal correction, apical derotation and thoracic kyphosis with similar length of fusion and better lumbar lordosis restoration.

An Evolution Gaining Momentum-The Growing Role of Artificial Intelligence in the Diagnosis and Treatment of Spinal Diseases.

In recent years, applications using artificial intelligence have been gaining importance in the diagnosis and treatment of spinal diseases. In our review, we describe the basic features of artificial intelligence which are currently applied in the field of spine diagnosis and treatment, and we provide an orientation of the recent technical developments and their applications. Furthermore, we point out the possible limitations and challenges in dealing with such technological advances. Despite the momentary limitations in practical application, artificial intelligence is gaining ground in the field of spine treatment. As an applying physician, it is therefore necessary to engage with it in order to benefit from those advances in the interest of the patient and to prevent these applications being misused by non-medical partners.

AI Prediction of Neuropathic Pain after Lumbar Disc Herniation-Machine Learning Reveals Influencing Factors.

The treatment options for neuropathic pain caused by lumbar disc herniation have been debated controversially in the literature. Whether surgical or conservative therapy makes more sense in individual cases can hardly be answered. We have investigated whether a machine learning-based prediction of outcome, regarding neuropathic pain development, after lumbar disc herniation treatment is possible. The extensive datasets of 123 consecutive patients were used to predict the development of neuropathic pain, measured by a visual analogue scale (VAS) for leg pain and the Oswestry Disability Index (ODI), at 6 weeks, 6 months and 1 year after treatment of lumbar disc herniation in a machine learning approach. Using a decision tree regressor algorithm, a prediction quality within the limits of the minimum clinically important difference for the VAS and ODI value could be achieved. An analysis of the influencing factors of the algorithm reveals the important role of psychological factors as well as body weight and age with pre-existing conditions for an accurate prediction of neuropathic pain. The machine learning algorithm developed here can enable an assessment of the course of treatment after lumbar disc herniation. The early, comparative individual prediction of a therapy outcome is important to avoid unnecessary surgical therapies as well as insufficient conservative therapies and prevent the chronification of neuropathic pain.

Directed invasion of cancer cell spheroids inside 3D collagen matrices oriented by microfluidic flow in experiment and simulation.

Invasion is strongly influenced by the mechanical properties of the extracellular matrix. Here, we use microfluidics to align fibers of a collagen matrix and study the influence of fiber orientation on invasion from a cancer cell spheroid. The microfluidic setup allows for highly oriented collagen fibers of tangential and radial orientation with respect to the spheroid, which can be described by finite element simulations. In invasion experiments, we observe a strong bias of invasion towards radial as compared to tangential fiber orientation. Simulations of the invasive behavior with a Brownian diffusion model suggest complete blockage of migration perpendicularly to fibers allowing for migration exclusively along fibers. This slows invasion toward areas with tangentially oriented fibers down, but does not prevent it.

Etiology, Risk Factors, and Diagnosis of Back Pain in Children and Adolescents: Evidence- and Consensus-Based Interdisciplinary Recommendations.

Using a structured approach and expert consensus, we developed an evidence-based guideline on the diagnosis of back pain and the treatment of non-specific back pain in children and adolescents. The first part comprises etiology, risk factors, and diagnosis. The second part, published in the same issue, includes treatment and prevention. A comprehensive and systematic literature search was conducted to identify relevant guidelines and studies. Based on the findings of this literature search, recommendations on risk factors and diagnosis were formulated and voted on by experts in a structured consensus-building process. Notable red flags for specific back pain and evidence-based risk factors for non-specific back pain in children and adolescents were identified. Only three evidence-based recommendations could be formulated for causes, red flags, and risk factors for back pain, while two recommendations are based on expert consensus. Regarding diagnostics, eight expert consensus recommendations and one evidence-based recommendation could be provided. Despite the importance of adequate diagnosis for the treatment of back pain in children and adolescents, results of this work confirm the deficit in research investment in this area.

Treatment of Unspecific Back Pain in Children and Adolescents: Results of an Evidence-Based Interdisciplinary Guideline.

Using a structured approach and expert consensus, we developed an evidence-based guideline on the treatment and prevention of non-specific back pain in children and adolescents. A comprehensive and systematic literature search identified relevant guidelines and studies. Based on the findings of this literature search, recommendations on treatment and prevention were formulated and voted on by experts in a structured consensus-building process. Physical therapy (particularly physical activity) and psychotherapy (particularly cognitive behavioral therapy) are recommended for treating pediatric non-specific back pain. Intensive interdisciplinary treatment programs should be provided for chronic and severe pain. Drug therapy should not be applied in children and adolescents. Further research on non-specific back pain in childhood and adolescence is strongly needed to reduce the imbalance between the high burden of non-specific back pain in childhood and adolescence and the low research activity in this field.

Anterior vertebroplasty of adjacent levels after vertebral body replacement.

The aim of this study was to evaluate the feasibility of a new method, which should help to avoid cage subsidence after vertebrectomy in short fusions. After implantation of an extendable vertebral body replacement (VBR) the two adjacent endplates to the fractured or destroyed vertebra were augmented with bone cement using the anterior approach in 20 patients with short circumferential fusion. All patients were followed up for 2 years clinically and radiographically. X-rays were reviewed for kyphosis, cage subsidence, presence of a solid fusion mass and instrumentation failure. Changes in every day activities (Oswestry Disability Index-ODI) and visual analogue scale (VAS), pain score, as well as technique-related complications were examined. The mean amount of kyphosis correction was 12.8° (±6.4°) and changed by only 0.3° (±0.4°) until last follow-up. Pain (VAS) and ODI scores were significantly improved and did not change until last follow-up. In all but two cases, the authors observed solid union with incorporation of the cage. No surgery-related complications were recorded. In one case revision was advised because of non-union due to septic loosening. The augmentation of the adjacent vertebras after anterior VBR provides an enhancement of the interface between cage and vertebra in cases with poor bone quality and in revision surgery. The technique is simple and safe, as the needles can be placed under visual control. Cement augmentation of the endplates may reduce interbody device subsidence.

Combined Artificial Intelligence Approaches Analyzing 1000 Conservative Patients with Back Pain-A Methodological Pathway to Predicting Treatment Efficacy and Diagnostic Groups.

Patients with back pain are common and present a challenge in everyday medical practice due to the multitude of possible causes and the individual effects of treatments. Predicting causes and therapy efficien cy with the help of artificial intelligence could improve and simplify the treatment. In an exemplary collective of 1000 conservatively treated back pain patients, it was investigated whether the prediction of therapy efficiency and the underlying diagnosis is possible by combining different artificial intelligence approaches. For this purpose, supervised and unsupervised artificial intelligence methods were analyzed and a methodology for combining the predictions was developed. Supervised AI is suitable for predicting therapy efficiency at the borderline of minimal clinical difference. Non-supervised AI can show patterns in the dataset. We can show that the identification of the underlying diagnostic groups only becomes possible through a combination of different AI approaches and the baseline data. The presented methodology for the combined application of artificial intelligence algorithms shows a transferable path to establish correlations in heterogeneous data sets when individual AI approaches only provide weak results.

Reduction of radiation dose during radiofrequency denervation of the lumbar facet joints using the new targeting system SabreSource: a prospective study in 20 patients.

BACKGROUND AND AIMS: Facet joint denervation is a frequently performed technique to treat facet joint syndrome. Most often this technique is used under fluoroscopic guidance implicating high radiation doses for both patients and surgeons. This prospective study was performed to evaluate the effectiveness in reducing radiation dose during radiofrequency ablation therapy of the lumbar facet joints and to evaluate the feasibility and possibilities of the new real time image guidance system SabreSource. MATERIALS AND METHODS: As much as 20 consecutive patients with radiofrequency ablation therapy of the facet joints L4 to S1 were included. Ten patients were treated by fluoroscopic control alone; the following 10 patients were treated with the SabreSource image guidance system. A total of 40 thermal ablations to the facet joints were performed. Each patient was given one thermal ablation on both sides of the vertebral segment, either to the facet joints of L4-L5 or of L5-S1. Pain, according to the visual analogue scale (VAS), was documented before and 6 h after the intervention. Radiation dose, time of radiation and the number of shots needed to place the radiofrequency cannula were recorded. RESULTS: No complications occurred. Before therapy, the mean VAS in all patients was 7.6 (range 6-10). After therapy the mean VAS in all patients was 3.4 (range 0-5). Compared to the fluoroscopy-guided thermal ablation therapy the SabreSource system significantly reduced the number of fluoroscopy exposures (reduction 23.53%, p = 0.02), the time of radiation exposure (reduction 21.2%, p = 0.03) and the mean entrance surface dose (reduction 30.46%, p = 0.01). CONCLUSION: The SabreSource System reduces radiation exposure and radiation dose in the radiofrequency denervation therapy of the lumbar facet joints and can be applied for other minimally invasive techniques.

Expression of TRAIL and the death receptors DR4 and DR5 correlates with progression of degeneration in human intervertebral disks.

Intervertebral disks degenerate far earlier than other musculoskeletal tissues and apoptosis has been suggested to have a vital function in promoting the degeneration process that is strongly associated with back pain. However, the molecular mediators of apoptosis in the intervertebral disk are poorly understood. Fas/FasL, TRAIL/DR4, TRAIL/DR5 and TNF-alpha/TNFR1 are ligand/receptor pairs of the tumor necrosis factor/nerve growth factor family, which are able to induce apoptosis by trimerization of the receptor by its corresponding ligand. We investigated which of these molecules are expressed in intervertebral disks and whether their expression correlates to disk degeneration. Intervertebral disks from 28 donors (age 12-70 years) suffering from scoliosis, vertebrae fracture or disk degeneration were scored histologically for degeneration and analyzed for gene expression of FasL/Fas, TRAIL/DR4, TNF-alpha/TNFR1 and caspase 8. Protein expression of FasL and TRAIL was assessed by immunohistology and apoptotic cell death was quantified by poly(ADP-ribose) polymerase (PARP) p85 staining. Isolated disk cells were analyzed by flow cytometry for Fas, FasL, TRAIL, DR4 and DR5 expression. Gene expression of TRAIL (P=0.002) and caspase 8 (P=0.027) significantly correlated with degeneration. TRAIL expression further correlated with cellularity (P=0.04), muccoid matrix changes (P=0.009) and tears and cleft formation (P=0.019). FasL and TRAIL expression was confirmed by immunohistology and PARP cleavage was significantly associated with degeneration (P=0.027). Flow cytometry on isolated disk cells revealed correlations between DR4 and degeneration (P=0.014), DR4/DR5 double-positive cells and degeneration (P=0.019), as well as DR5 and changes in tissue granularity (P=0.03). This is the first study that shows that intervertebral disk cells express TRAIL, DR4 and DR5, which correlate to the degenerative state of the disk. Therefore, disk cells inherit the molecular machinery to induce and undergo cellular apoptosis, and the frequency of cytokine expression suggests that the TRAIL/DR4/DR5 axis is an important molecular mediator of apoptosis induction in disk tissue.

Reduction of radiation dose during facet joint injection using the new image guidance system SabreSource: a prospective study in 60 patients.

Interventional procedures are associated with high radiation doses for both patients and surgeons. To reduce the risk from ionizing radiation, it is essential to minimize radiation dose. This prospective study was performed to evaluate the effectiveness in reducing radiation dose during facet joint injection in the lumbar spine and to evaluate the feasibility and possibilities of the new real time image guidance system SabreSource. A total of 60 patients, treated with a standardized injection therapy of the facet joints L4-L5 or L5-S1, were included in this study. A total of 30 patients were treated by fluoroscopy guidance alone, the following 30 patients were treated using the new SabreSource system. Thus a total of 120 injections to the facet joints were performed. Pain, according to the visual analogue scale (VAS), was documented before and 6 h after the intervention. Radiation dose, time of radiation and the number of exposures needed to place the needle were recorded. No significant differences concerning age (mean age 60.5 years, range 51-69), body mass index (mean BMI 26.2, range 22.2-29.9) and preoperative pain (VAS 7.9, range 6-10) were found between the two groups. There was no difference in pain reduction between the two groups (60 vs. 61.5%; P = 0.001) but the radiation dose was significantly smaller with the new SabreSource system (reduction of radiation dose 32.7%, P = 0.01; reduction of mean entrance surface dose 32.3%, P = 0.01). The SabreSource System significantly reduced the radiation dose received during the injection therapy of the lumbar facet joints. With minimal effort for the setup at the beginning of a session, the system is easy to handle and can be helpful for other injection therapies (e.g. nerve root block therapies).

Fiber stiffness, pore size and adhesion control migratory phenotype of MDA-MB-231 cells in collagen gels.

Cancer cell migration is influenced by cellular phenotype and behavior as well as by the mechanical and chemical properties of the environment. Furthermore, many cancer cells show plasticity of their phenotype and adapt it to the properties of the environment. Here, we study the influence of fiber stiffness, confinement, and adhesion properties on cancer cell migration in porous collagen gels. Collagen gels with soft fibers abrogate migration and promote a round, non-invasive phenotype. Stiffer collagen fibers are inherently more adhesive and lead to the existence of an adhesive phenotype and in general confined migration due to adhesion. Addition of TGF-ß lowers adhesion, eliminates the adhesive phenotype and increases the amount of highly motile amoeboid phenotypes. Highest migration speeds and longest displacements are achieved in stiff collagen fibers in pores of about cell size by amoeboid phenotypes. This elucidates the influence of the mechanical properties of collagen gels on phenotype and subsequently migration and shows that stiff fibers, cell sized pores, and low adhesion, are optimal conditions for an amoeboid phenotype and efficient migration.

Open versus minimally invasive TLIF: literature review and meta-analysis.

STUDY DESIGN: This study is a comparative, literature review. OBJECTIVE: The aim of this study is to provide a comparative analysis of open vs. minimally invasive TLIF using a literature review and a meta-analysis. Lumbar interbody fusion is a well-established surgical procedure for treating several spinal disorders. Transforaminal lumbar interbody fusion (TLIF) was initially introduced in the early 1980s. To reduce approach-related morbidity associated with traditional open TLIF (OTLIF), minimally invasive TLIF (MITLIF) was developed. We aimed to provide a comparative analysis of open vs. minimally invasive TLIF using a literature review. METHODS: We searched the online database PubMed (2005-2017), which yielded an initial 194 studies. We first searched the articles' abstracts. Based on our inclusion criteria, we excluded 162 studies and included 32 studies: 18 prospective, 13 retrospective, and a single randomized controlled trial. Operative time, blood loss, length of hospital stay, radiation exposure time, complication rate, and pain scores (visual analogue scale, Oswestry Disability Index) for both techniques were recorded and presented as means. We then performed a meta-analysis. RESULTS: The meta-analysis for all outcomes showed reduced blood loss (P < 0.00001) and length of hospital stay (P < 0.00001) for MITLIF compared with OTLIF, but with increased radiation exposure time with MITLIF (P < 0.00001). There was no significant difference in operative time between techniques (P = 0.78). The complication rate was lower with MITLIF (11.3%) vs. OTLIF (14.2%), but not statistically significantly different (P = 0.05). No significant differences were found in visual analogue scores (back and leg) and Oswestry Disability Index scores between techniques, at the final follow-up. CONCLUSION: MITLIF and OTLIF provide equivalent long-term clinical outcomes. MITLIF had less tissue injury, blood loss, and length of hospital stay. MITLIF is also a safe alternative in obese patients and, in experienced hands, can also be used safely in select cases of spondylodiscitis even with epidural abscess. MITLIF is also a cost-saving procedure associated with reduced hospital and social costs. Long-term studies are required to better evaluate controversial items such as operative time.

The effect of platelet-rich plasma on healing in critical-size long-bone defects.

The role of platelet-rich plasma (PRP) as a promoter of bone healing remains controversial. The hypothesis investigated was that PRP improves bone healing of a critical-size diaphyseal radius defect in a rabbit model. The bone defect was filled with a high-surface ceramic scaffold, calcium-deficient hydroxyapatite (CDHA), with the addition of allogenic PRP, mesenchymal stem cells (MSC) or both. PRP yielded better bone formation than the empty CDHA scaffold as determined by both histology and micro-computer tomography (p<0.05) after 16 weeks, whereas no difference was observed on biomechanical testing. Similar behavior was found in samples with MSC; however, the combination of MSC and PRP did not further improve bone healing. Furthermore, the resorption of CDHA was improved by the addition of PRP, MSC and MSC/PRP, but there were no differences between the groups. The areas of bone formation were greater in areas adjacent to the bone resection areas and towards the intact ulna. In conclusion, PRP improves bone healing in a diaphyseal rabbit model on CDHA and the combination of CDHA. This study supports the allogenic use of PRP for bone healing as an off-the-shelf therapy.